Organ mimic device with microchannels and methods of use and manufacturing thereof

ABSTRACT

System and method includes a body having a central microchannel separated by one or more porous membranes. The membranes are configured to divide the central microchannel into a two or more parallel central microchannels, wherein one or more first fluids are applied through the first central microchannel and one or more second fluids are applied through the second or more central microchannels. The surfaces of each porous membrane can be coated with cell adhesive molecules to support the attachment of cells and promote their organization into tissues on the upper and lower surface of the membrane. The pores may be large enough to only permit exchange of gases and small chemicals, or to permit migration and transchannel passage of large proteins and whole living cells. Fluid pressure, flow and channel geometry also may be varied to apply a desired mechanical force to one or both tissue layers.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation application of co-pending U.S. patentapplication Ser. No. 13/054,095 filed on Jan. 4, 2011, which applicationis a 371 National Phase Entry Application of International ApplicationNo. PCT/US2009/050830 filed Jul. 16, 2009, which designates the U.S.,and which claims the benefit of priority under 35 U.S.C. §119(e) of U.S.Provisional Application No. 61/081,080 filed Jul. 16, 2008; acontinuation application of co-pending U.S. patent application Ser. No.14/099,113 filed Dec. 6, 2013, which is a continuation application ofU.S. patent application Ser. No. 13/054,095 filed on Jan. 4, 2011, whichapplication is a 371 National Phase Entry Application of InternationalApplication No. PCT/US2009/050830 filed Jul. 16, 2009, which designatesthe U.S., and which claims the benefit of priority under 35 U.S.C.§119(e) of U.S. Provisional Application No. 61/081,080 filed Jul. 16,2008; a continuation application of co-pending U.S. patent applicationSer. No. 14/099,247, filed Dec. 6, 2013, which is a continuationapplication of U.S. patent application Ser. No. 13/054,095 filed on Jan.4, 2011, which application is a 371 National Phase Entry Application ofInternational Application No. PCT/US2009/050830 filed Jul. 16, 2009,which designates the U.S., and which claims the benefit of priorityunder 35 U.S.C. §119(e) of U.S. Provisional Application No. 61/081,080filed Jul. 16, 2008. The contents of all of which are incorporatedherein by reference in their entirety.

GOVERNMENT SUPPORT

This invention was made with Government support under Grant No.: NIH R01ES016665-01A1 awarded by the National Institutes of Health. TheGovernment has certain rights in the invention.

TECHNICAL FIELD

The present disclosure relates generally to an organ mimic device withmicrochannels and methods of use and manufacturing thereof.

BACKGROUND

Mechanical forces—pushes, pulls, tensions, compressions—are importantregulators of cell development and behavior. Tensegrity provides thestructure that determines how these physical forces are distributedinside a cell or tissue, and how and where they exert their influence.

In the human body, most cells are constantly subjected to mechanicalforces, such as tension or compression. Application of mechanical strainto cells in culture simulates the in vivo environment, causing dramaticmorphologic changes and biomechanical responses in the cells. There areboth long and short term changes that occur when cells are mechanicallyloaded in culture, such as alterations in the rate and amount of DNA orRNA synthesis or degradation, protein expression and secretion, the rateof cell division and alignment, changes in energy metabolism, changes inrates of macromolecular synthesis or degradation, and other changes inbiochemistry and bioenergetics.

Every cell has an internal scaffolding, or cytoskeleton, a latticeformed from molecular “struts and wires”. The “wires” are acrisscrossing network of fine cables, known as microfilaments, thatstretch from the cell membrane to the nucleus, exerting an inward pull.Opposing the pull are microtubules, the thicker compression-bearing“struts” of the cytoskeleton, and specialized receptor molecules on thecell's outer membrane that anchor the cell to the extracellular matrix,the fibrous substance that holds groups of cells together. This balanceof forces is the hallmark of tensegrity.

Tissues are built from groups of cells, like eggs sitting on the “eggcarton” of the extracellular matrix. The receptor molecules anchoringcells to the matrix, known as integrins, connect the cells to the widerworld. Mechanical force on a tissue is felt first by integrins at theseanchoring points, and then is carried by the cytoskeleton to regionsdeep inside each cell. Inside the cell, the force might vibrate orchange the shape of a protein molecule, triggering a biochemicalreaction, or tug on a chromosome in the nucleus, activating a gene.

Cells also can be said to have “tone,” just like muscles, because of theconstant pull of the cytoskeletal filaments. Much like a stretchedviolin string produces different sounds when force is applied atdifferent points along its length, the cell processes chemical signalsdifferently depending on how much it is distorted.

A growth factor will have different effects depending on how much thecell is stretched. Cells that are stretched and flattened, like those inthe surfaces of wounds, tend to grow and multiply, whereas roundedcells, cramped by overly crowded conditions, switch on a “suicide”program and die. In contrast, cells that are neither stretched norretracted carry on with their intended functions.

Another tenet of cellular tensegrity is that physical location matters.When regulatory molecules float around loose inside the cell, theiractivities are little affected by mechanical forces that act on the cellas a whole. But when they're attached to the cytoskeleton, they becomepart of the larger network, and are in a position to influence cellulardecision-making Many regulatory and signaling molecules are anchored onthe cytoskeleton at the cell's surface membrane, in spots known asadhesion sites, where integrins cluster. These prime locations are keysignal-processing centers, like nodes on a computer network, whereneighboring molecules can receive mechanical information from theoutside world and exchange signals.

Thus, assessing the full effects of drugs, drug delivery vehicles,nanodiagnostics or therapies or environmental stressors, such asparticles, gases, and toxins, in a physiological environment requiresnot only a study of the cell-cell and cell-chemical interactions, butalso a study of how these interactions are affected by physiologicalmechanical forces in both healthy tissues and tissues affected withdiseases.

Methods of altering the mechanical environment or response of cells inculture have included wounding cells by scraping a monolayer, applyingmagnetic or electric fields, or by applying static or cyclic tension orcompression with a screw device, hydraulic pressure, or weights directlyto the cultured cells. Shear stress has also been induced by subjectingthe cells to fluid flow. However, few of these procedures have allowedfor quantitation of the applied strains or provided regulation toachieve a broad reproducible range of cyclic deformations within aculture microenvironment that maintains physiologically relevanttissue-tissue interactions.

Living organs are three-dimensional vascularized structures composed oftwo or more closely apposed tissues that function collectively andtransport materials, cells and information across tissue-tissueinterfaces in the presence of dynamic mechanical forces, such as fluidshear and mechanical strain. Creation of microdevices containing livingcells that recreate these physiological tissue-tissue interfaces andpermit fluid flow and dynamic mechanical distortion would have greatvalue for study of complex organ functions, e.g., immune celltrafficking, nutrient absorption, infection, oxygen and carbon dioxideexchange, etc., and for drug screening, toxicology, diagnostics andtherapeutics.

The alveolar-capillary unit plays a vital role in the maintenance ofnormal physiological function of the lung as well as in the pathogenesisand progression of various pulmonary diseases. Because of the complexarchitecture of the lung, the small size of lung alveoli and theirsurrounding microvessels, and the dynamic mechanical motions of thisorgan, it is difficult to study this structure at the microscale.

The lung has an anatomically unique structure having a hierarchicalbranching network of conducting tubes that enable convective gastransport to and from the microscopic alveolar compartments where gasexchange occurs. The alveolus is the most important functional unit ofthe lung for normal respiration, and it is most clinically relevant inthat it is the blood-gas barrier or interface, as well as the site wheresurfactants act to permit air entry and where immune cells, pathogensand fluids accumulate in patients with acute respiratory distresssyndrome (ARDS) or infections, such as pneumonia.

The blood-gas barrier or tissue-tissue interface between the pulmonarycapillaries and the alveolar lumen is composed of a single layer ofalveolar epithelium closely juxtaposed to a single layer of capillaryendothelium separated by a thin extracellular matrix (ECM), which formsthrough cellular and molecular self-assembly in the embryo. Virtuallyall analysis of the function of the alveolar-capillary unit has beencarried out in whole animal studies because it has not been possible toregenerate this organ-level structure in vitro.

A major challenge lies in the lack of experimental tools that canpromote assembly of multi-cellular and multi-tissue organ-likestructures that exhibit the key structural organization, physiologicalfunctions, and physiological or pathological mechanical activity of thelung alveolar-capillary unit, which normally undergoes repeatedexpansion and contraction during each respiratory cycle. This limitationcould be overcome if it were possible to regenerate this organ-levelstructure and recreate its physiological mechanical microenvironment invitro. However, this has not been fully accomplished.

What is needed is a organ mimic device capable of being used in vitro orin vivo which performs tissue-tissue related functions and which alsoallows cells to naturally organize in the device in response to not onlychemical but also mechanical forces and allows the study of cellbehavior through a membrane which mimics tissue-tissue physiology.

OVERVIEW

System and method comprises a body having a central microchannelseparated by one or more porous membranes. The membranes are configuredto divide the central microchannel into a two or more closely apposedparallel central microchannels, wherein one or more first fluids areapplied through the first central microchannel and one or more secondfluids are applied through the second or more central microchannels. Thesurfaces of each porous membrane can be coated with cell adhesivemolecules to support the attachment of cells and promote theirorganization into tissues on the upper and lower surface of eachmembrane, thereby creating one or more tissue-tissue interfacesseparated by porous membranes between the adjacent parallel fluidchannels. The membrane may be porous, flexible, elastic, or acombination thereof with pores large enough to only permit exchange ofgases and small chemicals, or large enough to permit migration andtranschannel passage of large proteins, as well as whole living cells.Fluid pressure, flow characteristics and channel geometry also may bevaried to apply a desired fluid shear stress to one or both tissuelayers.

In an embodiment, operating channels adjacent to the central channel areapplied a positive or negative pressure which creates a pressuredifferential that causes the membrane to selectively expand and retractin response to the pressure, thereby further physiologically simulatingmechanical force of a living tissue-tissue interface.

In another embodiment, three or more parallel microchannels areseparated by a plurality of parallel porous membranes which are lined bya common tissue type in the central channel and two different tissuetypes on the opposite sides of the membranes in the two outer channels.An example would be a cancer mimic device in which cancer cells aregrown in the central micro channel and on the inner surfaces of bothporous membranes, while capillary endothelium is grown on the oppositesurface of one porous membrane and lymphatic endothelium is grown on theopposite surface of the second porous membrane. This recreates the tumormicroarchitecture and permits study of delivery of oxygen, nutrients,drugs and immune cells via the vascular conduit as well as tumor cellegress and metastasis via the lymphatic micro channel.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings, which are incorporated into and constitute apart of this specification, illustrate one or more examples ofembodiments and, together with the description of example embodiments,serve to explain the principles and implementations of the embodiments.In the drawings:

FIG. 1 illustrates a block diagram of a system employing an exampleorgan mimic device in accordance with an embodiment.

FIG. 2A illustrates a perspective view of a organ mimic device inaccordance with an embodiment.

FIG. 2B illustrates an exploded view of the organ mimic device inaccordance with an embodiment.

FIGS. 2C-2D illustrate perspective views of tissue-tissue interfaceregions of the device in accordance with an embodiment.

FIGS. 2E-2G illustrate top down cross sectional views of thetissue-tissue interface regions of the device in accordance with one ormore embodiments.

FIGS. 3A-3B illustrate perspective views of tissue-tissue interfaceregions of the device in accordance with an embodiment.

FIGS. 3C-3E illustrate perspective views of the membrane in accordancewith one or more embodiments.

FIGS. 4A-4C illustrate perspective views of the formation of themembrane of a two channel device in accordance with an embodiment.

FIG. 4D illustrates a side view of the membrane of the tissue-tissueinterface device in accordance with an embodiment.

FIGS. 5A-5E illustrate perspective views of the formation of the organmimic device in accordance with an embodiment.

FIG. 6 illustrates a system diagram employing an organ mimic device withmultiple channels in accordance with an embodiment.

FIGS. 7A-7B illustrate perspective views of the organ mimic device inaccordance with an embodiment.

FIG. 7C illustrates a side view of the membrane of the organ mimicdevice in accordance with an embodiment.

FIGS. 8 and 9 illustrate ROS generation over time in accordance with anexperiment conducting with the present device.

DESCRIPTION OF EXAMPLE EMBODIMENTS

Example embodiments are described herein in the context of an organsimulating device and methods of use and manufacturing thereof. Those ofordinary skill in the art will realize that the following description isillustrative only and is not intended to be in any way limiting. Otherembodiments will readily suggest themselves to such skilled personshaving the benefit of this disclosure. Reference will now be made indetail to implementations of the example embodiments as illustrated inthe accompanying drawings. The same reference indicators will be usedthroughout the drawings and the following description to refer to thesame or like items. It is understood that the phrase “an embodiment”encompasses more than one embodiment and is thus not limited to only oneembodiment for brevity's sake.

In accordance with this disclosure, the organ mimic device (alsoreferred to as “present device”) is preferably utilized in an overallsystem incorporating sensors, computers, displays and other computingequipment utilizing software, data components, process steps and/or datastructures. The components, process steps, and/or data structuresdescribed herein with respect to the computer system with which theorgan mimic device is employed may be implemented using various types ofoperating systems, computing platforms, computer programs, and/orgeneral purpose machines. In addition, those of ordinary skill in theart will recognize that devices of a less general purpose nature, suchas hardwired devices, field programmable gate arrays (FPGAs),application specific integrated circuits (ASICs), or the like, may alsobe used without departing from the scope and spirit of the inventiveconcepts disclosed herein.

Where a method comprising a series of process steps is implemented by acomputer or a machine with use with the organ mimic device describedbelow and those process steps can be stored as a series of instructionsreadable by the machine, they may be stored on a tangible medium such asa computer memory device (e.g., ROM (Read Only Memory), PROM(Programmable Read Only Memory), EEPROM (Electrically EraseableProgrammable Read Only Memory), FLASH Memory, Jump Drive, and the like),magnetic storage medium (e.g., tape, magnetic disk drive, and the like),optical storage medium (e.g., CD-ROM, DVD-ROM, paper card, paper tapeand the like) and other types of program memory.

Embodiments of the present device can be applied in numerous fieldsincluding basic biological science, life science research, drugdiscovery and development, drug safety testing, chemical and biologicalassays, as well as tissue and organ engineering. In an embodiment, theorgan mimic device can be used as microvascular network structures forbasic research in cardiovascular, cancer, and organ-specific diseasebiology. Furthermore, one or more embodiments of the device findapplication in organ assist devices for liver, kidney, lung, intestine,bone marrow, and other organs and tissues, as well as in organreplacement structures.

The cellular responses to the various environmental cues can bemonitored using various systems that can be combined with the presentdevice. One can monitor changes in pH using well known sensors. One canalso sample cells, continuously or periodically for measurement ofchanges in gene transcription or changes in cellular biochemistry orstructural organization. For example, one can measure reactive oxygenspecies (ROIs) that are a sign of cellular stress. One can also subjectthe “tissue” grown on the porous membrane to microscopic analysis,immunohistochemical analysis, in situ hybridization analysis, or typicalpathological analysis using staining, such as hematoxylin and eosinstaining Samples for these analysis can be carried out in real-time, ortaken after an experiment or by taking small biopsies at differentstages during a study or an experiment.

One can subject the cells grown on the membrane to other cells, such asimmune system cells or bacterial cells, to antibodies orantibody-directed cells, for example to target specific cellularreceptors. One can expose the cells to viruses or other particles. Toassist in detection of movement of externally supplied substances, suchas cells, viruses, particles or proteins, one can naturally label themusing typical means such as radioactive or fluorescent labels.

Cells can be grown, cultured and analyzed using the present device for1, 2, 3, 4, 5, 6, or 7 days, between at least 1-2 weeks, and even over 2weeks. For example, as discussed below, it has been shown thatco-culture of alveolar epithelial cells with pulmonary microvascularendothelial cells on a thin porous membrane in an embodiment of thedescribed device could be grown for over two weeks without loss ofviability of the cells.

The organ mimic device described herein has many different applicationsincluding, but not limited to, identification of markers of disease;assessing efficacy of anti-cancer therapeutics; testing gene therapyvectors; drug development; screening; studies of cells, especially stemcells and bone marrow cells; studies on biotransformation, absorption,clearance, metabolism, and activation of xenobiotics; studies onbioavailability and transport of chemical or biological agents acrossepithelial or endothelial layers; studies on transport of biological orchemical agents across the blood-brain barrier; studies on transport ofbiological or chemical agents across the intestinal epithelial barrier;studies on acute basal toxicity of chemical agents; studies on acutelocal or acute organ-specific toxicity of chemical agents; studies onchronic basal toxicity of chemical agents; studies on chronic local orchronic organ-specific toxicity of chemical agents; studies onteratogenicity of chemical agents; studies on genotoxicity,carcinogenicity, and mutagenicity of chemical agents; detection ofinfectious biological agents and biological weapons; detection ofharmful chemical agents and chemical weapons; studies on infectiousdiseases; studies on the efficacy of chemical or biological agents totreat disease; studies on the optimal dose range of agents to treatdisease; prediction of the response of organs in vivo to biologicalagents; prediction of the pharmacokinetics of chemical or biologicalagents; prediction of the pharmacodynamics of chemical or biologicalagents; studies concerning the impact of genetic content on response toagents; studies on gene transcription in response to chemical orbiological agents; studies on protein expression in response to chemicalor biological agents; studies on changes in metabolism in response tochemical or biological agents. The organ mimic device can also be usedto screen on the cells, for an effect of the cells on the materials (forexample, in a manner equivalent to tissue metabolism of a drug).

The present device may be used by one to simulate the mechanical loadenvironment of walking, running, breathing, peristalsis, flow of flow orurine, or the beat of a heart, to cells cultured from mechanicallyactive tissues, such as heart, lung, skeletal muscle, bone, ligament,tendon, cartilage, smooth muscle cells, intestine, kidney, endothelialcells and cells from other tissues. Rather than test the biological orbiochemical responses of a cell in a static environment, theinvestigator can apply a range of frequencies, amplitudes and durationof mechanical stresses, including tension, compression and shear, tocultured cells.

A skilled artisan can implant various types of cells on the surfaces ofthe membrane. Cells include any cell type from a multicellularstructure, including nematodes, amoebas, up to mammals such as humans.Cell types implanted on the device depend on the type of organ or organfunction one wishes to mimic, and the tissues that comprise thoseorgans. More details of the various types of cells implantable on themembrane of the present device are discussed below.

One can also co-culture various stem cells, such as bone marrow cells,induced adult stem cells, embryonal stem cells or stem cells isolatedfrom adult tissues on either or both sides of the porous membrane. Usingdifferent culture media in the chambers feeding each layer of cells, onecan allow different differentiation cues to reach the stem cell layersthereby differentiating the cells to different cell types. One can alsomix cell types on the same side of the membrane to create co-cultures ofdifferent cells without membrane separation.

Using the organ mimic device described herein, one can studybiotransformation, absorption, clearance, metabolism, and activation ofxenobiotics, as well as drug delivery. The bioavailability and transportof chemical and biological agents across epithelial layers as in theintestine, endothelial layers as in blood vessels, and across theblood-brain barrier can also be studied. The acute basal toxicity, acutelocal toxicity or acute organ-specific toxicity, teratogenicity,genotoxicity, carcinogenicity, and mutagenicity, of chemical agents canalso be studied. Effects of infectious biological agents, biologicalweapons, harmful chemical agents and chemical weapons can also bedetected and studied. Infectious diseases and the efficacy of chemicaland biological agents to treat these diseases, as well as optimal dosageranges for these agents, can be studied. The response of organs in vivoto chemical and biological agents, and the pharmacokinetics andpharmacodynamics of these agents can be detected and studied using thepresent device. The impact of genetic content on response to the agentscan be studied. The amount of protein and gene expression in response tochemical or biological agents can be determined. Changes in metabolismin response to chemical or biological agents can be studied as wellusing the present device.

The advantages of the organ mimic device, as opposed to conventionalcell cultures or tissue cultures, are numerous. For instance, when cellsare placed in the organ mimic device, fibroblast, SMC (smooth musclecell) and EC (endothelial cell) differentiation can occur thatreestablishes a defined three-dimensional architectural tissue-tissuerelationships that are close to the in vivo situation, and cellfunctions and responses to pharmacological agents or active substancesor products can be investigated at the tissue and organ levels.

In addition, many cellular or tissue activities are amenable todetection in the organ mimic device, including, but not limited to,diffusion rate of the drugs into and through the layered tissues intransported flow channel; cell morphology, differentiation and secretionchanges at different layers; cell locomotion, growth, apoptosis, and thelike. Further, the effect of various drugs on different types of cellslocated at different layers of the system may be assessed easily.

For drug discovery, for example, there can be two advantages for usingthe organ mimic device described herein: (1) the organ mimic device isbetter able to mimic in vivo layered architecture of tissues andtherefore allow one to study drug effect at the organ level in additionto at the cellular and tissue levels; and (2) the organ mimic devicedecreases the use of in vivo tissue models and the use of animals fordrug selection and toxicology studies.

In addition to drug discovery and development, the organ mimic devicedescribed herein may be also useful in basic and clinical research. Forexample, the organ mimic device can be used to research the mechanism oftumorigenesis. It is well established that in vivo cancer progression ismodulated by the host and the tumor micro-environment, including thestromal cells and extracellular matrix (ECM). For example, stromal cellswere found being able to convert benign epithelial cells to malignantcells, thereby ECM was found to affect the tumor formation. There isgrowing evidence that cells growing in defined architecture are moreresistant to cytotoxic agents than cells in mono layers. Therefore, aorgan mimic device is a better means for simulating the original growthcharacteristics of cancer cells and thereby better reflects the realdrug's sensitivity of in vivo tumors.

The organ mimic device can be employed in engineering a variety oftissues including, but not limited to, the cardiovascular system, lung,intestine, kidney, brain, bone marrow, bones, teeth, and skin. If thedevice is fabricated with a suitable biocompatible and/or biodegradablematerial, such as poly-lactide-co-glycolide acid (PLGA), the organ mimicdevice may be used for transplantation or implantation in vivo.Moreover, the ability to spatially localize and control interactions ofseveral cell types presents an opportunity to engineer hierarchically,and to create more physiologically correct tissue and organ analogs. Thearrangement of multiple cell types in defined arrangement has beneficialeffects on cell differentiation, maintenance, and functional longevity.

The organ mimic device can also allow different growth factors,chemicals, gases and nutrients to be added to different cell typesaccording to the needs of cells and their existence in vivo. Controllingthe location of those factors or proteins may direct the process ofspecific cell remodeling and functioning, and also may provide themolecular cues to the whole system, resulting in such beneficialdevelopments as neotissue, cell remodeling, enhanced secretion, and thelike.

In yet another aspect, the organ mimic device can be utilized as multicell type cellular microarrays, such as microfluidic devices. Using theorgan mimic device, pattern integrity of cellular arrays can bemaintained. These cellular microarrays may constitute the future“lab-on-a-chip”, particularly when multiplexed and automated. Theseminiaturized multi cell type cultures will facilitate the observation ofcell dynamics with faster, less noisy assays, having built-in complexitythat will allow cells to exhibit in vivo-like responses to the array.

In yet another aspect, the organ mimic device can be utilized asbiological sensors. Cell-based biosensors can provide more informationthan other biosensors because cells often have multifacetedphysiological responses to stimuli, as well as novel mechanisms toamplify these responses. All cell types in the organ mimic device can beused to monitor different aspects of an analyte at the same time;different cell type in the organ mimic device can be used to monitordifferent analytes at the same time; or a mixture of both types ofmonitoring. Cells ranging from E. coli to cells of mammalian lines havebeen used as sensors for applications in environmental monitoring, toxindetection, and physiological monitoring.

In yet another aspect, the organ mimic device can be used inunderstanding fundamental processes in cell biology and cell-ECMinteractions. The in vivo remodeling process is a complicated, dynamic,reciprocal process between cells and ECMs. The organ mimic device wouldbe able to capture the complexity of these biological systems, renderingthese systems amenable to investigation and beneficial manipulation.Furthermore, coupled with imaging tools, such as fluorescencemicroscopy, microfluorimetry or optical coherence tomography (OCT),real-time analysis of cellular behavior in the multilayered tissues isexpected using the device. Examples of cell and tissue studies amenableto real-time analysis include cell secretion and signaling, cell-cellinteractions, tissue-tissue interactions, dynamic engineered tissueconstruction and monitoring, structure-function investigations in tissueengineering, and the process of cell remodeling matrices in vitro.

Another example of the use of this device is to induce tissue-tissueinterfaces and complex organ structures to form within the device byimplanting it in vivo within the body of a living animal, and allowingcells and tissues to impregnate the device and establish normaltissue-tissue interfaces. Then the whole device and contained cells andtissues is surgically removed while perfusing it through one or more ofthe fluid channels with medium and gases necessary for cell survival.This complex organ mimic may then be maintained viable in vitro throughcontinuous perfusion and used to study highly complex cell and tissuefunctions in their normal 3D context with a level of complexity notpossible using any existing in vitro model system.

In particular, a microchannel device may be implanted subcutaneously invivo into an animal in which the device contains bone-inducingmaterials, such as demineralized bone powder or bone morphogenicproteins (BMPs), in a channel that has one or more corresponding portsopen to the surrounding tissue space. The second channel would be closedduring implantation by closing its end ports or filling it with a solidremovable materials, such as a solid rod. As a result of wound healing,connective tissues containing microcapillaries and mesenchymal stemcells would grow into the open channels of the device and, due to thepresence of the bone-inducing material, will form bone with spaces thatrecruit circulating hematopoietic precursor cells to form fullyfunctional bone marrow, as shown in past studies.

Once this process is complete, the surgical site would be reopened, andthe second channel would be reopened by removing the rod or plugs andwould then be connected with catheters linked to a fluid reservoir sothat culture medium containing nutrients and gases necessary for cellsurvival could be pumped through the second channel and passed throughthe pores of the membrane into the first channel containing the formedbone marrow. The entire microchannel device could then be cut free fromthe surrounding tissue, and with the medium flowing continuously intothe device, would be removed from the animal and passed to a tissueculture incubator and maintained in culture with continuous fluid flowthrough the second channel, and additional flow can be added to thefirst channel as well if desired by connecting to their inlet and outletports. The microchannel device would then be used to study intact bonemarrow function in vitro as in a controlled environment.

Both biocompatible and biodegradable materials can be used in thepresent device to facilitate in vivo implantation of the present device.One can also use biocompatible and biodegradable coatings. For example,one can use ceramic coatings on a metallic substrate. But any type ofcoating material and the coating can be made of different types ofmaterials: metals, ceramics, polymers, hydrogels or a combination of anyof these materials.

Biocompatible materials include, but are not limited to an oxide, aphosphate, a carbonate, a nitride or a carbonitride. Among the oxide thefollowing ones are preferred: tantalum oxide, aluminum oxide, iridiumoxide, zirconium oxide or titanium oxide. In some cases the coating canalso be made of a biodegradable material that will dissolve over timeand may be replaced by the living tissue. Substrates are made ofmaterials such as metals, ceramics, polymers or a combination of any ofthese. Metals such as stainless steel, Nitinol, titanium, titaniumalloys, or aluminum and ceramics such as zirconia, alumina, or calciumphosphate are of particular interest.

The biocompatible material can also be biodegradable in that it willdissolve over time and may be replaced by the living tissue. Suchbiodegradable materials include, but are not limited to, poly(lacticacid-co-glycolic acid), polylactic acid, polyglycolic acid (PGA),collagen or other ECM molecules, other connective tissue proteins,magnesium alloys, polycaprolactone, hyaluric acid, adhesive proteins,biodegradable polymers, synthetic, biocompatible and biodegradablematerial, such as biopolymers, bioglasses, bioceramics, calcium sulfate,calcium phosphate such as, for example, monocalcium phosphatemonohydrate, monocalcium phosphate anhydrous, dicalcium phosphatedihydrate, dicalcium phosphate anhydrous, tetracalcium phosphate,calcium orthophosphate phosphate, calcium pyrophosphate,alpha-tricalcium phosphate, beta-tricalcium phosphate, apatite such ashydroxyapatite, or polymers such as, for example,poly(alpha-hydroxyesters), poly(ortho esters), poly(ether esters),polyanhydrides, poly(phosphazenes), polypropylene fumarates), poly(esteramides), poly(ethylene fumarates), poly(amino acids), polysaccharides,polypeptides, poly(hydroxy butyrates), poly(hydroxy valerates),polyurethanes, poly(malic acid), polylactides, polyglycolides,polycaprolactones, poly(glycolide-co-trimethylene carbonates),polydioxanones, or copolymers, terpolymers thereof or blends of thosepolymers, or a combination of biocompatible and biodegradable materials.One can also use biodegradable glass and bioactive glass self-reinforcedand ultrahigh strength bioabsorbable composites assembled from partiallycrystalline bioabsorbable polymers, like polyglycolides, polylactidesand/or glycolide/lactide copolymers.

These materials preferably have high initial strength, appropriatemodulus and strength retention time from 4 weeks up to 1 year in vivo,depending on the implant geometry. Reinforcing elements such as fibersof crystalline polymers, fibers of carbon in polymeric resins, andparticulate fillers, e.g., hydroxyapatite, may also be used to providethe dimensional stability and mechanical properties of biodegradabledevices. The use of interpenetrating networks (IPN) in biodegradablematerial construction has been demonstrated as a means to improvemechanical strength. To further improve the mechanical properties ofIPN-reinforced biodegradable materials, the present device may beprepared as semi-interpenetrating networks (SIPN) of crosslinkedpolypropylene fumarate within a host matrix ofpoly(lactide-co-glycolide) 85:15 (PLGA) orpoly(l-lactide-co-d,l-lactide) 70:30 (PLA) using different crosslinkingagents. One can also use natural poly(hydroxybutyrate-co-9%hydroxyvalerate) copolyester membranes as described in Charles-HilaireRivard et al. (Journal of Applied Biomaterials, Volume 6 Issue 1, Pages65-68, 1 Sep. 2004). A skilled artisan will be able to also select otherbiodegradable materials suitable for any specific purposes and cell andtissue types according to the applications in which the device is used.

The device as described can also be used as therapeutic devices, whenplaced in vivo. One can re-create organ mimics, such as bone marrow orlymph nodes by placing the devices in, for example an animal modelallowing the device to be inhabited by living cells and tissues, andthen removing the entire device with living cells while perfusing thevascular channel with medium. The device can then be removed and keptalive ex vivo for in vitro or ex vivo studies. In particular, themembrane may be coated with one or more cell layers on at least one sideof the membrane in vitro. In this embodiment, the cells are platedoutside an organism. In an embodiment, the membrane is coated with oneor more cell layers on at least one side of the membrane in vivo. Onecan treat one side of the membrane in vitro and the other side in vivo.One can also have one or both sides initially coated with one cell typein vitro and then implant the device to attract additional cell layersin vivo.

In general, the present disclosure is directed to device and method ofuse in which the device includes a body having a central microchannelseparated by one or more porous membranes. The membrane(s) areconfigured to divide the central microchannel into two or more closelyapposed parallel central microchannels, wherein one or more first fluidsare applied through the first central microchannel and one or moresecond fluids are applied through the second or more centralmicrochannels. The surfaces of each porous membrane can be coated withcell adhesive molecules to support the attachment of cells and promotetheir organization into tissues on the upper and lower surface of themembrane, thereby creating a tissue-tissue interface separated by aporous membrane between the adjacent parallel fluid channels. Themembrane may be porous, flexible, elastic, or a combination thereof withpores large enough to only permit exchange of gases and small chemicals,or large enough to permit migration and transchannel passage of largeproteins, and whole living cells. Fluid pressure, flow and channelgeometry also may be varied to apply a desired fluid shear stress to oneor both tissue layers.

In a non-limiting example embodiment, the device is configured to mimicoperation of a lung, whereby lung epithelium cells self assemble on onesurface of the ECM membrane and lung capillary endothelium cells selfassemble on the opposite face of the same porous membrane. The devicethereby allows simulation of the structure and function of a functionalalveolar-capillary unit that can be exposed to physiological mechanicalstrain to simulate breathing or to both air-borne and blood-bornechemical, molecular, particulate and cellular stimuli to investigate theexchange of chemicals, molecules, and cells across this tissue-tissueinterface through the pores of the membrane. The device impacts thedevelopment of in vitro lung models that mimic organ-level responseswhich are able to be analyzed under physiological and pathologicalconditions. This system may be used in several applications including,but not limited to, drug screening, drug delivery, vaccine delivery,biodetection, toxicology, physiology and organ/tissue engineeringapplications.

FIG. 1 illustrates a block diagram of the overall system employing theinventive device in accordance with an embodiment. As shown in FIG. 1,the system 100 includes an organ mimic device 102, one or more fluidsources 104, 104 _(N) coupled to the device 102, one or more optionalpumps 106 coupled to the fluid source 104 and device 102. One or moreCPUs 110 are coupled to the pump 106 and preferably control the flow offluid in and out of the device 102. The CPU preferably includes one orprocessors 112 and one or more local/remote storage memories 114. Adisplay 116 is coupled to the CPU 110, and one or more pressure sources118 are coupled to the CPU 110 and the device 102. The CPU 110preferably controls the flow and rate of pressurized fluid to thedevice. It should be noted that although one interface device 102 isshown and described herein, it is contemplated that a plurality ofinterface devices 102 may be tested and analyzed within the system 100as discussed below.

As will be discussed in more detail, the organ mimic device 102preferably includes two or more ports which place the microchannels ofthe device 102 in communication with the external components of thesystem, such as the fluid and pressure sources. In particular, thedevice 102 is coupled to the one or more fluid sources 104 _(N) in whichthe fluid source may contain air, blood, water, cells, compounds,particulates, and/or any other media which are to be delivered to thedevice 102. In an embodiment, the fluid source 104 provides fluid to oneor more microchannels of the device 102 and also preferably receives thefluid which exits the device 102. It is contemplated that the fluidexiting the device 102 may additionally or alternatively be collected ina fluid collector or reservoir 108 separate from the fluid source 104.Thus, it is possible that separate fluid sources 104, 104 _(N)respectively provide fluid to and remove fluid from the device 102.

In an embodiment, fluid exiting the device 102 may be reused andreintroduced into the same or different input port through which itpreviously entered. For example, the device 102 may be set up such thatfluid passed through a particular central microchannel is recirculatedback to the device and is again run through the same centralmicrochannel. This could be used, for instance, to increase theconcentration of an analyte in the fluid as it is recirculated thedevice. In another example, the device 102 may be set up such that fluidpassed through the device and is recirculated back into the device andthen subsequently run through another central microchannel. This couldbe used to change the concentration or makeup of the fluid as it iscirculated through another microchannel.

One or more pumps 106 are preferably utilized to pump the fluid into thedevice 102, although pumps in general are optional to the system. Fluidpumps are well known in the art and are not discussed in detail herein.As will be discussed in more detail below, each microchannel portion ispreferably in communication with its respective inlet and/or outletport, whereby each microchannel portion of allow fluid to flowtherethrough.

Each microchannel in the device preferably has dedicated inlet andoutlet ports which are connected to respective dedicated fluid sourcesand/or fluid collectors to allow the flow rates, flow contents,pressures, temperatures and other characteristics of the media to beindependently controlled through each central microchannel. Thus, onecan also monitor the effects of various stimuli to each of the cell ortissue layers separately by sampling the separate fluid channels for thedesired cellular marker, such as changes in gene expression at RNA orprotein level.

The cell injector/remover 108 component is shown in communication withthe device 102, whereby the injector/remover 108 is configured toinject, remove and/or manipulate cells, such as but not limited toepithelial and endothelial cells, on one or more surfaces of theinterface membrane within the device 102 independent of cells introducedinto the device via the inlet port(s) 210, 218. For example, bloodcontaining magnetic particles which pull pathogenic cells may becultured in a separate device whereby the mixture can be laterintroduced into the system via the injector at a desired time withouthaving to run the mixture through the fluid source 104. In anembodiment, the cell injector/remover 108 is independently controlled,although the injector/remover 108 may be controlled by the CPU 110 asshown in FIG. 1. The cell injector/remover 108 is an optional componentand is not necessary.

Although not required, pressure may be applied from the one or morepressure sources 118 to create a pressure differential to causemechanical movements within the device 102. In an embodiment in whichpressures are used with the device, the pressure source 118 iscontrolled by the CPU 110 to apply a pressure differential within thedevice to effectively cause one or more membranes (FIGS. 3A-3B) withinthe device to expand and/or contract in response to the applied pressuredifferential. In an embodiment, the pressure applied to the device 100by the pressure source 118 is a positive pressure, depending on theconfiguration or application of the device. Additionally oralternatively, the pressure applied by the pressure source 118 is anegative pressure, such as vacuum or suction, depending on theconfiguration or application of the device. The pressure source 118 ispreferably controlled by the CPU 110 to apply pressure at set timedintervals or frequencies to the device 102, whereby the timing intervalsmay be set to be uniform or non-uniform. The pressure source 118 may becontrolled to apply uniform pressure in the timing intervals or mayapply different pressures at different intervals. For instance, thepressure applied by the pressure source 118 may have a large magnitudeand/or be set at a desired frequency to mimic a person running orundergoing exertion. The pressure source 118 may also apply slow,irregular patterns, such as simulating a person sleeping. In anembodiment, the CPU 110 operates the pressure source 118 to randomlyvary intervals of applying pressure to cause cyclic stretching patternsto simulate irregularity in breath rate and tidal volumes during naturalbreathing.

One or more sensors 120 may be coupled to the device 102 to monitor oneor more areas within the device 102, whereby the sensors 120 providemonitoring data to the CPU 110. One type of sensor 120 is preferably apressure sensor which provides data regarding the amount of pressure inone or more operating or central microchannels the device 102. Pressuredata from opposing sides of the microchannel walls may be used tocalculate real-time pressure differential information between theoperating and central microchannels. The monitoring data would be usedby the CPU 110 to provide information on the device's operationalconditions as well as how the cells are behaving within the device 102in particular environments in real time. The sensor 120 may be anelectrode, have infrared, optical (e.g. camera, LED), or magneticcapabilities or utilize any other appropriate type of technology toprovide the monitoring data. For instance, the sensor may be one or moremicroelectrodes which analyze electrical characteristics across themembrane (e.g. potential difference, resistance, and short circuitcurrent) to confirm the formation of an organized barrier, as well asits fluid/ion transport function across the membrane. It should be notedthat the sensor 120 may be external to the device 102 or be integratedwithin the device 102. It is contemplated that the CPU 110 controlsoperation of the sensor 120, although it is not necessary. The data ispreferably shown on the display 116.

FIG. 2A illustrates a perspective view of the tissue interface device inaccordance with an embodiment. In particular, as shown in FIG. 2A, thedevice 200 (also referred to reference numeral 102) preferably includesa body 202 having a branched microchannel design 203 in accordance withan embodiment. The body 202 may be made of a flexible material, althoughit is contemplated that the body be alternatively made of a non-flexiblematerial. It should be noted that the microchannel design 203 is onlyexemplary and not limited to the configuration shown in FIG. 2A. Thebody 202 is preferably made of a flexible biocompatible polymer,including but not limited to, polydimethyl siloxane (PDMS), orpolyimide. It is also contemplated that the body 202 may be made ofnon-flexible materials like glass, silicon, hard plastic, and the like.Although it is preferred that the interface membrane be made of the samematerial as the body 202, it is contemplated that the interface membranebe made of a material that is different than the body of the device.

The device in FIG. 2A includes a plurality of ports 205 which will bedescribed in more detail below. In addition, the branched configuration203 includes a tissue-tissue interface simulation region (membrane 208in FIG. 2B) where cell behavior and/or passage of gases, chemicals,molecules, particulates and cells are monitored. FIG. 2B illustrates anexploded view of the organ mimic device in accordance with anembodiment. In particular, the outer body 202 of the device 200 ispreferably comprised of a first outer body portion 204, a second outerbody portion 206 and an intermediary porous membrane 208 configured tobe mounted between the first and second outer body portions 204, 206when the portions 204, 206 are mounted to one another to form theoverall body.

FIG. 2B illustrates an exploded view of the device in accordance with anembodiment. As shown in FIG. 2B, the first outer body portion 204includes one or more inlet fluid ports 210 preferably in communicationwith one or more corresponding inlet apertures 211 located on an outersurface of the body 202. The device 100 is preferably connected to thefluid source 104 via the inlet aperture 211 in which fluid travels fromthe fluid source 104 into the device 100 through the inlet fluid port210.

Additionally, the first outer body portion 204 includes one or moreoutlet fluid ports 212 preferably in communication with one or morecorresponding outlet apertures 215 on the outer surface of the body 202.In particular, fluid passing through the device 100 exits the device 100to a fluid collector 108 or other appropriate component via thecorresponding outlet aperture 215. It should be noted that the device200 may be set up such that the fluid port 210 is an outlet and fluidport 212 is an inlet. Although the inlet and outlet apertures 211, 215are shown on the top surface of the body 202, one or more of theapertures may be located on one or more sides of the body.

In an embodiment, the inlet fluid port 210 and the outlet fluid port 212are in communication with the first central microchannel 250A (see FIG.3A) such that fluid can dynamically travel from the inlet fluid port 210to the outlet fluid port 212 via the first central microchannel 250A,independently of the second central microchannel 250B (see FIG. 3A).

It is also contemplated that the fluid passing between the inlet andoutlet fluid ports may be shared between the central sections 250A and250B. In either embodiment, characteristics of the fluid flow, such asflow rate and the like, passing through the central microchannel 250A iscontrollable independently of fluid flow characteristics through thecentral microchannel 250B and vice versa.

In addition, the first portion 204 includes one or more pressure inletports 214 and one or more pressure outlet ports 216 in which the inletports 214 are in communication with corresponding apertures 217 locatedon the outer surface of the device 100. Although the inlet and outletapertures are shown on the top surface of the body 202, one or more ofthe apertures may alternatively be located on one or more sides of thebody.

In operation, one or more pressure tubes (not shown) connected to thepressure source 118 (FIG. 1) provides positive or negative pressure tothe device via the apertures 217. Additionally, pressure tubes (notshown) are connected to the device 100 to remove the pressurized fluidfrom the outlet port 216 via the apertures 223. It should be noted thatthe device 200 may be set up such that the pressure port 214 is anoutlet and pressure port 216 is an inlet. It should be noted thatalthough the pressure apertures 217, 223 are shown on the top surface ofthe body 202, it is contemplated that one or more of the pressureapertures 217, 223 may be located on one or more side surfaces of thebody 202.

Referring to FIG. 2B, the second outer body portion 206 preferablyincludes one or more inlet fluid ports 218 and one or more outlet fluidports 220. It is preferred that the inlet fluid port 218 is incommunication with aperture 219 and outlet fluid port 220 is incommunication with aperture 221, whereby the apertures 219 and 221 arepreferably located on the outer surface of the second outer body portion206. Although the inlet and outlet apertures are shown on the surface ofthe body 202, one or more of the apertures may be alternatively locatedon one or more sides of the body.

As with the first outer body portion 204 described above, one or morefluid tubes connected to the fluid source 104 (FIG. 1) are preferablycoupled to the aperture 219 to provide fluid to the device 100 via port218. Additionally, fluid exits the device 100 via the outlet port 220and out aperture 221 to a fluid reservoir/collector 108 or othercomponent. It should be noted that the device 200 may be set up suchthat the fluid port 218 is an outlet and fluid port 220 is an inlet.

In addition, it is preferred that the second outer body portion 206includes one or more pressure inlet ports 222 and one or more pressureoutlet ports 224. In particular, it is preferred that the pressure inletports 222 are in communication with apertures 227 and pressure outletports 224 are in communication with apertures 229, whereby apertures 227and 229 are preferably located on the outer surface of the secondportion 206. Although the inlet and outlet apertures are shown on thebottom surface of the body 202, one or more of the apertures may bealternatively located on one or more sides of the body. Pressure tubesconnected to the pressure source 118 (FIG. 1) are preferably engagedwith ports 222 and 224 via corresponding apertures 227 and 229. Itshould be noted that the device 200 may be set up such that the pressureport 222 is an outlet and fluid port 224 is an inlet.

In an embodiment, the membrane 208 is mounted between the first portion204 and the second portion 206, whereby the membrane 208 is locatedwithin the body 202 of the device 200 (see FIG. 5E). In an embodiment,the membrane 208 is a made of a material having a plurality of pores orapertures therethrough, whereby molecules, cells, fluid or any media iscapable of passing through the membrane 208 via one or more pores in themembrane 208. As discussed in more detail below, it is contemplated inan embodiment that the porous membrane 208 may be made of a materialwhich allows the membrane 208 to undergo stress and/or strain inresponse to pressure differentials present between the centralmicrochannels 250A, 250B and the operating microchannels. Alternatively,the porous membrane 208 is relatively inelastic in which the membrane208 undergoes minimal or no movement while media is passed through oneor more of the central microchannels 250A, 250B and cells organize andmove between the central microchannels 250A, 250B via the porousmembrane.

Referring FIG. 2C illustrates a perspective view of the tissue-tissueinterface region of the first outer portion 204 of the body taken atline C-C (from FIG. 2B). As shown in FIG. 2C, the top portion of thetissue-tissue interface region 207A is within the body of the firstportion 204 and includes a top portion of a central microchannel 230 andone or more top portion side operating microchannels 232 locatedadjacent to the central microchannel 230. Microchannel walls 234preferably separate the central microchannel 230 from the operatingmicrochannels 232 such that fluid traveling through the centralmicrochannel 230 does not pass into operating microchannels 232.Likewise, the channel walls 234 prevent pressurized fluid passing alongoperating microchannels 232 from entering the central microchannel 230.It should be noted that a pair of operating microchannels 232 are shownon opposing sides of central microchannel 230 in FIGS. 2C and 3A,however it is contemplated that the device may incorporate more than twooperating microchannels 232. It is also contemplated that the device 200may include only one operating microchannel 232 adjacent to the centralmicrochannel 230.

FIG. 2D illustrates a perspective view of the tissue interface regiontaken at line D-D of the second outer portion 206 of the body. As shownin FIG. 2D, the tissue interface region includes a bottom portion of thecentral microchannel 240 and at least two bottom portions of operatingmicrochannels 242 located adjacent to the central microchannel 240portion. A pair of channel walls 234 preferably separate the centralmicrochannel 240 from the operating microchannels 232 such that fluidtraveling through the central microchannel 230 does not pass intooperating microchannels 232. Likewise, the channel walls 234 preventpressurized fluid passing along operating microchannels 232 fromentering the central microchannel 230.

As shown in FIGS. 2C and 2D, the top and bottom portions 230 and 240 ofthe central microchannel each have a range of width dimension (shown asB) between 50 and 1000 microns, and preferably around 400 microns. Itshould be noted that other width dimensions are contemplated dependingon the type of physiological system which is being mimicked in thedevice. Additionally, the top and bottom portions of the operatingmicrochannels 232 and 242 each have a width dimension (shown as A)between 25 and 800 microns, and preferably around 200 microns, althoughother width dimensions are contemplated. The height dimensions of thecentral and/or operating microchannels range between 50 microns andseveral centimeters, and preferably around 200 microns. The microchannelwalls 234, 244 preferably have a thickness range between 5 microns to 50microns, although other width dimensions are contemplated depending onthe material used for the walls, application in which the device is usedand the like.

FIG. 3A illustrates a perspective view of the tissue interface regionwithin the body in accordance with an embodiment. In particular, FIG. 3Aillustrates the first portion 207A and the second portion 207B mated toone another whereby the side walls 228 and 238 as well as channel walls234, 244 form the overall central microchannel 250 and operatingmicrochannels 252. As stated above, it is preferred that centralmicrochannel 250 and operating microchannels 252 are separated by thewalls 234, 244 such that fluid is not able to pass between the channels250, 252.

The membrane 208 is preferably positioned in the center of the centralmicrochannel 250 and is oriented along a plane parallel to the x-y planeshown in FIG. 3A. It should be noted that although one membrane 208 isshown in the central microchannel 250, more than one membrane 208 may beconfigured within the central microchannel 250, as discussed in moredetail below. In addition to being positioned within the centralmicrochannel 250, the membrane 208 is sandwiched in place by channelwalls 234, 244 during formation of the device.

The membrane 208 preferably separates the overall central microchannel250 into two or more distinct central microchannels 250A and 250B. Itshould be noted that although the membrane 208 is shown midway throughthe central microchannel 250, the membrane 208 may alternatively bepositioned vertically off-center within the central microchannel 250,thus making one of the central microchannel sections 250A, 250B largerin volume or cross-section than the other microchannel section.

As will be discussed in more detail below, the membrane 208 may have atleast a portion which is porous to allow cells or molecules to passtherethrough. Additionally or alternatively, at least a portion of themembrane 208 may have elastic or ductile properties which allow themembrane 208 to be manipulated to expand/contract along one or moreplanar axe. Thus, it is contemplated that one or more portions of themembrane 208 may be porous and elastic or porous, but inelastic.

With regard to the porous and elastic membrane, a pressure differentialmay be applied within the device to cause relative continuous expansionand contraction of the membrane 208 along the x-y plane. In particular,as stated above, one or more pressure sources preferably applypressurized fluid (e.g. air) along the one or more operatingmicrochannels 252, whereby the pressurized fluid in the microchannels252 creates a pressure differential on the microchannel walls 234, 244.The membrane 208 may have an elasticity depending on the type ofmaterial that it is made of. If the membrane 208 is made of more thanone material, the weight ratio of the respective materials which make upthe membrane is a factor in determining the elasticity. For example, inthe embodiment that the membrane 208 is made of PDMS, the Young'smodulus values are in the ranges of 12 kPa-20 MPa, although otherelasticity values are contemplated.

In the embodiments shown in FIGS. 3A and 3B, the pressurized fluid is avacuum or suction force that is applied only through the operatingmicrochannels 252. The difference in pressure caused by the suctionforce against the microchannel walls 234, 244 causes the walls 234, 244to bend or bulge outward toward the sides of the device 228, 238 (seeFIG. 3B). Considering that the membrane 208 is mounted to and sandwichedbetween the walls 234, 244, the relative movement of the walls 234, 244thereby causes the opposing ends of the membrane to move along with thewalls to stretch (shown as 208′ in FIG. 3B) along the membrane's plane.This stretching mimics the mechanical forces experienced by atissue-tissue interface, for example, in the lung alveolus duringbreathing, and thus provides the important regulation for cellular selfassembly into tissue structures and cell behavior.

When the negative pressure is no longer applied (and/or positivepressure is applied to the operating channels), the pressuredifferential between the operating channels 252 and the central channel250 decreases and the channel walls 234, 244 retract elastically towardtheir neutral position (as in FIG. 3A). During operation, the negativepressure is alternately applied in timed intervals to the device 200 tocause continuous expansion and contraction of the membrane 208 along itsplane, thereby mimicking operation of the tissue-tissue interface of theliving organ within a controlled in vitro environment. As will bediscussed, this mimicked organ operation within the controlledenvironment allows monitoring of cell behavior in the tissues, as wellas passage of molecules, chemicals, particulates and cells with respectto the membrane and the associated first and second microchannels 250A,250B.

It should be noted that the term pressure differential in the presentspecification relates to a difference of pressure on opposing sides of aparticular wall between the central microchannel and the outer operatingchannel. It is contemplated that the pressure differential may becreated in a number of ways to achieve the goal of expansion and/orcontraction of the membrane 208. As stated above, a negative pressure(i.e. suction or vacuum) may be applied to one or more of the operatingchannels 252. Alternatively, it is contemplated that the membrane 208 ispre-loaded or pre-stressed to be in an expanded state by default suchthat the walls 234, 244 are already in the bent configuration, as showin FIG. 3B. In this embodiment, positive pressure applied to theoperating channel 252 will create the pressure differential which causesthe walls 234, 244 to move inward toward the central microchannel (seein FIG. 3A) to contract the membrane 208.

It is also contemplated, in another embodiment, that a combination ofpositive and negative pressure is applied to one or more operatingmicrochannels 252 to cause movement of the membrane 208 along its planein the central microchannel. In any of the above embodiments, it isdesired that the pressure of the fluid in the one or more operatingchannels 252 be such that a pressure differential is in fact createdwith respect to the pressure of the fluid(s) in one or more of thecentral microchannel(s) 250A, 250B to cause relativeexpansion/contraction of the membrane 208. For example, fluid may have acertain pressure may be applied within the top central microchannel250A, whereby fluid in the bottom central microchannel 250B may have adifferent pressure. In this example, pressure applied to the one or moreoperating channels 252 must take into account the pressure of the fluidin either or both of the central microchannels 250A, 250B to ensuredesired expansion/contraction of the membrane 208.

It is possible, in an embodiment, for a pressure differential to existbetween the top and bottom microchannels 250A, 250B to cause at least aportion of the membrane 208 to expand and/or contract vertically in thez-direction in addition to expansion/contraction along the x-y plane.

In an embodiment, the expansion and retraction of the membrane 208preferably applies mechanical forces to the adherent cells and ECM thatmimic physiological mechanical cues that can influence transport ofchemicals, molecules particulates, and/or fluids or gas across thetissue-tissue interface, and alter cell physiology. It should be notedthat although pressure differentials created in the device preferablycause expansion/contraction of the membrane, it is contemplated thatmechanical means, such as micromotors or actuators, may be employed toassist or substitute for the pressure differential to causeexpansion/contraction of the cells on the membrane to modulatecellphysiology.

FIGS. 3E and 4C illustrate perspectives view of the membrane 208 whichincludes a plurality of apertures 302 extending therethrough inaccordance with an embodiment. In particular, the membrane shown inFIGS. 3E and 4C includes one or more of integrated pores or apertures302 which extend between a top surface 304 and a bottom surface 306 ofthe membrane 208.

The membrane is configured to allow cells, particulates, chemicalsand/or media to migrate between the central microchannel portions 250A,250B via the membrane 208 from one section of the central microchannelto the other or vice versa. The pore apertures are shown to have apentagonal cross sectional shape in FIGS. 4A-4C, although any othercross sectional shape is contemplated, including but not limited to, acircular shaped 302, hexagonal 308, square, elliptical 310 and the like.The pores 302, 308, 310 (generally referred to as reference numeral 302)preferably extend vertically between the top and bottom surfaces 304,306, although it is contemplated that they may extend laterally as wellbetween the top and bottom surfaces, as with pore 312. It should also benoted that the porous may additionally/alternatively incorporate slitsor other shaped apertures along at least a portion of the membrane 208which allow cells, particulates, chemicals and/or fluids to pass throughthe membrane 208 from one section of the central microchannel to theother.

The width dimension of the pores are preferably in the range of 0.5microns and 20 microns, although it is preferred that the widthdimension be approximately 10 microns. It is contemplated, however, thatthe width dimension be outside of the range provided above. In someembodiments, the membrane 208 has pores or apertures larger thantraditional molecular/chemical filtration devices, which allow cells aswell as molecules to migrate across the membrane 208 from onemicrochannel section (e.g. 250A) to the other microchannel section (e.g.250B) or vice versa. This may be useful in culturing cells whichpolarize in the top and bottom central channels in response to being inthe microchannel environment provided by the device whereby fluid(s) andcells are dynamically passed through pores that connect thesemicrochannels 250A, 250B.

As shown in FIG. 4B, the thickness of the membrane 208 may be between 70nanometers and 50 microns, although a preferred range of thickness wouldbetween 5 and 15 microns. It is also contemplated that the membrane 208be designed to include regions which have lesser or greater thicknessesthan other regions in the membrane 208. As shown in FIG. 3C, themembrane 208 is shown to have one or more decreased thickness areas 209relative to the other areas of the membrane 208. The decreased thicknessarea(s) 209 may run along the entire length or width of the membrane 208or may alternatively be located at only certain locations of themembrane 208. It should be noted that although the decreased thicknessarea 209 is shown along the bottom surface of the membrane 208 (i.e.facing microchannel 250B), it is contemplated that the decreasedthickness area(s) 209 may additionally/alternatively be on the opposingsurface of the membrane 208 (i.e. facing microchannel 250A). It shouldalso be noted that at least portions of the membrane 208 may have one ormore larger thickness areas 209′ relative to the rest of the membrane,as shown in FIG. 3D and capable of having the same alternatives as thedecreased thickness areas described above.

In an embodiment, the porous membrane 208 may be designed or surfacepatterned to include micro and/or nanoscopic patterns therein such asgrooves and ridges, whereby any parameter or characteristic of thepatterns may be designed to desired sizes, shapes, thicknesses, fillingmaterials, and the like.

In an embodiment, the membrane 208 is made of polydimethylsiloxane(PDMS) or any other polymeric compound or material, although this is notnecessary. For instance, the membrane 208 may be made of polyimide,polyester, polycarbonate, cyclicolefin copolymer,polymethylmethacrylate, nylon, polyisoprene, polybutadiene,polychlorophene, polyisobutylene, poly(styrene-butadiene-styrene),nitriles, the polyurethanes and the polysilicones. GE RTV 615, avinyl-silane crosslinked (type) silicone elastomer (family) may be used.Polydimethysiloxane (PDMS) membranes are available HT-6135 and HT-6240membranes from Bisco Silicons (Elk Grove, Ill.) and are useful inselected applications. The choice of materials typically depends uponthe particular material properties (e.g., solvent resistance, stiffness,gas permeability, and/or temperature stability) required for theapplication being conducted. Additional elastomeric materials that canbe used in the manufacture of the components of the microfluidic devicesdescribed in Unger et al., (2000 Science 288:113-116). Some elastomersof the present devices are used as diaphragms and in addition to theirstretch and relax properties, are also selected for their porosity,impermeability, chemical resistance, and their wetting and passivatingcharacteristics. Other elastomers are selected for their thermalconductivity. Micronics Parker Chomerics Thermagap material61-02-0404-F574 (0.020″ thick) is a soft elastomer (<5Shore A) needingonly a pressure of 5 to 10 psi to provide a thermal conductivity of 1.6W/m-° K. Deformable films, lacking elasticity, can also be used in themicrofluidic device. One may also use silk, ECM gels with or withoutcrosslinking as other such suitable materials to make the devices andmembranes as described.

It should be noted that although the central and operating microchannels250, 252 are shown to have substantially square or rectangular crosssections, other cross-sectional shapes are contemplated such ascircular, oval, hexagonal, and the like. It is also contemplated thatthe device 200 may have more or less than two operating channels 252and/or more or less than two central microchannels 250A, 250B inaccordance with an embodiment.

In an embodiment, it is contemplated that the central microchannel has anon-uniform width dimension B along at least a portion of its length inthe device. FIG. 2E illustrates a cross sectional top-down view of thetissue interface region 400 in accordance with an embodiment. As shownin FIG. 2E, the interface 400 includes the central microchannel 402along with adjacent operating channels 406 separated by microchannelwalls 404. In the embodiment in FIG. 2E, the central microchannel 402 isshown to have a gradually increasing width from width dimension C (atend 408) to width dimension D (at end 410). In the embodiment in FIG.2E, the operating channels 406 each have a correspondingly decreasingwidth dimension (from width dimension E at end 408 to width dimension Fat end 410). In another embodiment, as shown in FIG. 2F, the operatingchannels 406′ have a substantially uniform width dimension F at ends 408and 410. It is contemplated that the membrane (not shown) be placedabove the central microchannel 402 and mounted to the top surface of thewalls 404, whereby the membrane has a tapered shape similar to thecentral microchannel 402. The tapered membrane would thereby undergonon-uniform stretching in the direction of the arrows when the pressuredifferential is applied between the operating microchannels 406 and thecentral microchannel 402.

In another embodiment, the central microchannel may have a portion whichhas a partially circular cross sectional shape, as shown in FIG. 2G. Inparticular to the embodiment in FIG. 2G, a pressure differential createdbetween the central microchannel 502 and the adjacent operatingmicrochannels 504 will cause the microchannel walls 506 to move in thedirection represented by the arrows. With regard to the circular portion508 of the central microchannel 502, equiaxial outward movement of thewalls (as shown by the arrows) at the central portion 508 producesequiaxial stretching of the membrane (not shown) mounted atop of thewalls 506.

The device 200 described herein has potential for several applications.For example, in one application, the membrane 208 may be subjected tophysiological mechanical strain generated by cyclic stretching of themembrane 208 and/or the flow of biological fluids (e.g. air, mucus,blood) to recapitulate the native microenvironment of the alveoli andunderlying pulmonary capillaries. In an embodiment, the cultureconditions of cells upon the membrane 208 may be optimized underextracellular matrix (ECM) coating, media perfusion, or cyclicmechanical strain to maintain morphological and functionalcharacteristics of the co-cultured cells and to permit their directcellular interaction across the membrane 208. The device 200 would thuspermit long-term cell culture and dynamic mechanical stretching of aadjacent monolayers of lung epithelial or endothelial cells grown on themembrane at the same time.

In utilizing the membrane 208 in simulating the tissue-tissue interfacebetween the alveolar epithelium and pulmonary endothelium in the lung,one method may be to apply type I alveolar epithelial cells to the sideof the membrane 208 facing the first section 250A (hereinafter top sideof membrane) to mimic the epithelial layer. It is possible, however, tomix type I-like and type II-like alveolar epithelial cells at a ratio ofapproximately 7:13 to reconstruct the in vivo cellular composition ofthe alveolar epithelium. In the example method, lung microvascularendothelial cells are cultured on the opposite side of the membrane 208facing the second section 250B (hereinafter bottom side of membrane). Inthe example method, negative pressure is cyclically applied to thedevice 200 to cause the membrane 208 to continuously expand and contractalong its plane.

During such operation, a physiological alveolar-capillary unit may beformed on the membrane 208 since typical junctional structures may formon the membrane 207 and fluids as well as ions be transported across themembrane 208 between the first and second sections 250A, 250B. Theformation of tight junctions on the membrane 208 may be evaluated usingon-chip immunohistochemical detection of tight junction proteins such asZO-1 and occludin. Additionally or alternately, the exclusion offluorescently labeled large molecules (e.g. dextrans of differentweight) may be quantitated to determine the permeability of the membraneand optimize epithelial membrane barrier formation by varying cultureconditions. Additionally, histological, biochemical, andmicrofluorimetric techniques may be employed to demonstrate formation ofa functional alveolar-capillary unit that reproduces the key structuralorganization of its in vivo counterpart on the membrane 208.

In an example, the gas exchange function of the tissue-tissue interfaceself assembled on membrane 208 may be determined by injecting differentfluids, each having their own oxygen partial pressures and blood, intothe respective first and second sections 250A, 250B, whereby the firstsection 250A acts as the alveolar compartment and the second section250B acts as the microvascular compartment. A blood-gas measurementdevice preferably within the device 200 is used to measure the level ofoxygen in the blood in the respective sections 250A, 250B before andafter the passing of the blood through the device. For example, bloodcan flow through the channel 250B while air is being injected into theupper channel 250A, whereby the exiting air is collected and measured todetermine the oxygen level using an oximeter. Oximeters can beintegrated with the existing system or as a separate unit connected tothe outlet port of one or more central microchannels. In an embodiment,air or another medium with aerosols containing drugs or particulates mayflow through the device, whereby the transport of these drugs orparticulates to the blood via the membrane is then measured. It is alsocontemplated that pathogens or cytokines are added to the air or gaseousmedium side and then the sticking of immune cells to nearby capillaryendothelium and their passage along with edema fluid from the blood sideto the airway side, as well as pathogen entry into blood, are measured.

Since the functionality of an epithelium requires polarization ofconstituent cells, the structure of the membrane may be visualized usingtransmission electron microscopy, immunohistocytochemistry, confocalmicroscopy, or other appropriate means to monitor the polarization ofthe alveolar epithelial cell side of the membrane 208. In a lung mimicembodiment, a flourescent dye may be applied to the first and/or secondmicrochannels 250A, 250B to determine pulmonary surfactant production bythe airway epithelium at the membrane 208. In particular, alveolarepithelial cells on the membrane 208 can be monitored by measuring thefluorescence resulting from cellular uptake of the fluorescence dye thatspecifically labels intracellular storage of pulmonary surfactant (e.g.quinacrine) or using specific antibodies.

One of the unique capabilities of the tissue interface device 200 allowsdevelopment of in vitro models that simulate inflammatory responses ofthe lung at the organ level to allow study of how immune cells migratefrom the blood, through the endothelium and into the alveolarcompartment. One way this is achieved is by controlled and programmablemicrofluidic delivery of pro-inflammatory factors (e.g. IL-1β, TNF-α,IL-8, silica micro- and nanoparticles, pathogens) to the first section250A as well as whole human blood flowing or medium containingcirculating immune cells in the second section 250B. Electricalresistance and short circuit current across the membrane may bemonitored to study changes in the vascular permeability, extravasationof fluid and cell passage into the alveolar space during inflammatoryresponses. Fluorescence microscopy can be used to visualize dynamic cellmotile behavior during the extravasation response.

The tissue interface device 200 may also be used to examine hownanomaterials behave with respect to the lung tissue-tissue interface.In particular, nanomaterials (e.g. silica nanoparticles,superparamagnetic nanoparticles, gold nanoparticles, single-walledcarbon nanotubes) may be applied to the airway surface of the membrane208 to investigate potential toxic effects of nanomaterials on airway orendothelial cells grown on the membrane 208, as well as their passagefrom the airway channel into the blood channel. For instance, sensors120 can be used to monitor transmigration of nanomaterials throughtissue barriers formed on the membrane 208 and nanomaterial-inducedchanges in barrier functions such as gas exchange and fluid/iontransport.

The tissue interface device 200 permits direct analysis of a variety ofimportant areas of lung biology and physiology including but not limitedto gas exchange, fluid/ion transport, inflammation, infection,edema/respiratory distress syndrome, cancer and metastasis development,fungal infection, drug delivery as well as drug screening, biodetection,and pulmonary mechanotransduction. In addition, the device 200 allowsfor accurately modeling biological tissue-tissue interfaces found inother physiological systems such as the blood-brain barrier, intestine,bone marrow, glomerulus, and cancerous tumor microenvironment. As statedabove, more than one tissue interface device 200 may be multiplexed andautomated to provide high-throughput analysis of cell and tissueresponses to drugs, chemicals, particulates, toxins, pathogens or otherenvironmental stimuli for drug, toxin and vaccine screening, as well astoxicology and biodetection applications. The device may be used forstudying complex tissue and organ physiology in vitro, as well as tissueand organ engineering in vivo with biocompatible or biodegradeabledevices.

In an embodiment, the tissue interface device 200 can be used to produceartificial tissue layers therein. In the embodiment, two or moredifferent types of cells are applied on opposing surfaces of themembrane 208 and grown under conditions that mimic the appropriatephysiological environments. Additionally or alternatively, a pressuredifferential can be applied between the central microchannel and atleast one of the operating microchannels which causes the microchannelwalls to move and thus causes the membrane 208 to undergoexpansion/contraction along its plane.

In another example, the device 200 utilizes the porous membrane 208,whereby lung cells are grown on one side of the membrane 208 andendothelial cells are maintained on the other side of the membrane 208.During the operation of the device 200, these two cells layerscommunicate with each other through passage of chemical and molecularcues through the pores on the membrane 208. This communication may bemonitored and analyzed to understand how the cells function differentlyas a tissue-tissue interface, with or without physiological mechanicalsimulation, and compared to when grown as single tissue types inisolation as in standard tissue culture systems. By monitoring changesin cell and tissue physiology, as well as passage of chemicals,molecules, particulates and cells across this tissue-tissue interface,information is obtained which may be used to produce more effectivedrugs or therapies, to identify previously unknown toxicities, and tosignificantly shorten the timescale of these development processes. Inparticular, the behavior of cells in such a controlled environmentallows researchers to study a variety of physiological phenomena takingplace in the systems mentioned above that can not be recreated usingconventional in vitro culture techniques. In other words, the device 200functions to create a monitorable artificial blood-air barrier outside apatient's body and in a controllable environment that still retains keyphysiological functions and structures of the lung. It should be notedthat although the device above is described in terms of mimicking lungfunction, the device may easily be configured to mimic otherphysiological systems such as peristalsis and absorption in thegastrointestinal tract containing living microbial populations,perfusion and urine production in the kidney, function of theblood-brain barrier, effects of mechanical deformation on skin aging,bone marrow-microvessel interface with hematopoietic stem cell niche,and the like.

Details of membrane surface treatment and types of media which can beapplied to the membrane and/or through the central microchannels 250A,250B in operating the device will now be discussed. The membraneincluding the porous membrane can be coated with substances such asvarious cell adhesion promoting substances or ECM proteins, such asfibronectin, laminin or various collagen types or combinations thereof,as shown in FIG. 4D. In general, as shown in FIG. 4D, one or moresubstances 608 is shown on one surface of the membrane 604 whereasanother substance 610 is applied to the opposing surface of the membrane604, or both surfaces can be coated with the same substance. In someembodiments, the ECMs, which may be ECMs produced by cells, such asprimary cells or embryonic stem cells, and other compositions of matterare produced in a serum-free environment.

In an embodiment, one coats the membrane with a combination of a celladhesion factor and a positively-charged molecule that are bound to themembrane to improve cell attachment and stabilize cell growth. Thepositively charged molecule can be selected from the group consisting ofpolylysine, chitosan, poly(ethyleneimine) or acrylics polymerized fromacrylamide or methacrylamide and incorporating positively-charged groupsin the form of primary, secondary or tertiary amines, or quaternarysalts. The cell adhesion factor can be added to the membrane and ispreferably fibronectin, laminin, collagen, vitronectin or tenascin, orfragments or analogs having a cell binding domain thereof. Thepositively-charged molecule and the cell adhesion factor can becovalently bound to the membrane. In another embodiment, thepositively-charged molecule and the cell adhesion factor are covalentlybound to one another and either the positively-charged molecule or thecell adhesion factor is covalently bound to the membrane. Also, thepositively-charged molecule or the cell adhesion factor or both cam beprovided in the form of a stable coating non-covalently bound to themembrane.

In an embodiment, the cell attachment-promoting substances,matrix-forming formulations, and other compositions of matter aresterilized to prevent unwanted contamination. Sterilization may beaccomplished, for example, by ultraviolet light, filtration, or heat.Antibiotics may also be added, particularly during incubation, toprevent the growth of bacteria, fungi and other undesiredmicro-organisms. Such antibiotics include, by way of non-limitingexample, gentamicin, streptomycin, penicillin, amphotericin andciprofloxacin.

In another embodiment, the membrane is coated with cell cultures,including without limitation, primary cell cultures, established celllines, or stem cell cultures, such as ESC, attached to ECM substancesthat comprise or consist essentially of fibronectin or collagen.

In an embodiment, the primary cells or cell lines attached to themembrane may be alveolar cells, endothelial cells, intestinal cells,keratinocytes, which include without limitation, human dermalkeratinocytes, or any other type of cell listed elsewhere in thisspecification or well known to one skilled in the art. In otherembodiments, the primary cells may be fibroblast cells, which includewithout limitation, human fetal fibroblast cells. In some embodiments,the stem cells of the stem cell cultures are embryonic stem cells. Thesource of embryonic stem cells can include without limitation mammals,including non-human primates and humans. Non-limiting examples of humanembryonic stem cells include lines BG01, BG02, BG03, BG01v, CHA-hES-1,CHA-hES-2, FCNCBS1, FCNCBS2, FCNCBS3, H1, H7, H9, H13, H14, HSF-1, H9.1,H9.2, HES-1, HES-2, HES-3, HES-4, HES-5, HES-6, hES-1-2, hES-3-0,hES-4-0, hES-5-1, hES-8-1, hES-8-2, hES-9-1, hES-9-2, hES-101, hICM8,hICM9, hICM40, hICM41, hICM42, hICM43, HSF-6, HUES-1, HUES-2, HUES-3,HUES-4 HUES-5, HUES-6, HUES-7 HUES-8, HUES-9, HUES-10, HUES-11, HUES-12,HUES-13, HUES-14, HUESS-15, HUES-16, HUES-17, 13, 14, 16, 13.2, 13.3,16.2, J3, J3.2, MB01, MB02, MB03, Miz-hES1, RCM-1, RLS ES 05, RLS ES 07,RLS ES 10, RLS ES 13, RLS ES 15, RLS ES 20, RLS ES 21, SA01, SA02, andSA03. In an embodiment, the stem cells of the stem cell cultures areinduced pluripotent stem cells.

In an embodiment, the cell cultures may be cell cultures such as primarycell cultures or stem cell cultures which are serum-free. In some theseembodiments, a serum-free primary cell ECM is used in conjunction with aprimary cell serum-free medium (SFM). Suitable SFM include withoutlimitation (a) EPILIFE® Serum Free Culture Medium supplemented withEPILIFE® Defined Growth Supplement and (b) Defined Keratinocyte-SFMsupplemented with Defined Keratinocyte-SFM Growth Supplement, allcommercially available from Gibco/Invitrogen (Carlsbad, Calif., US). Insome of these embodiments, a serum-free stem cell ECM is used inconjunction with stem cell SFM. Suitable SFM include without limitationSTEMPRO® hESC Serum Free Media (SFM) supplemented with basic fibroblastgrowth factor and .beta.-mercaptoethanol, KNOCKOUT™ D-MEM supplementedwith KNOCKOUT™ Serum Replacement (SR), STEMPRO® MSC SFM and STEMPRO® NSCSFM, all commercially available from Gibco/Invitrogen (Carlsbad, Calif.,US).

In an embodiment, the compositions can also be xeno-free. A compositionof matter is said to be “xeno-free” when it is devoid of substances fromany animal other than the species of animal from which the cells arederived. In an embodiment, the cell cultures which may be cell culturessuch as primary cell cultures or stem cell cultures are xeno-free. Inthese embodiments, a xeno-free ECM which may be an ECM such as a primarycell ECM or ECM designed specifically to support stem cell growth ordifferentiation. These matrices may be specifically designed to bexeno-free.

In an embodiment, the cell cultures are primary cells or stem cellscultured in a conditioned culture medium. In other embodiments, theculture medium is an unconditioned culture medium.

In an embodiment, the cell culture conditions are completely defined. Inthese embodiments, one uses a completely defined cell culture medium inthe fluid chambers. Suitable media include without limitation, forprimary cells, EPILIFE® Serum Free Culture Medium supplemented withEPILIFE®. Defined Growth Supplement, and, for stem cells, STEMPRO® hESCSFM, all commercially available from Gibco/Invitrogen, Carlsbad, Calif.,US.

To study the effects of pharmaceuticals, environmental stressors,pathogens, toxins and such, one can add these into the desired cellculture medium suitable for growing the cells attached to the porousmembrane in the channel. Thus, one can introduce pathogens, such asbacteria, viruses, aerosols, various types of nanoparticles, toxins,gaseous substances, and such into the culture medium which flows in thechambers to feed the cells.

A skilled artisan will also be able to control the pH balance of themedium according to the metabolic activity of the cells to maintain thepH in a suitable level for any cell or tissue type in question. Monitorsand adjustment systems to monitor and adjust pH may be inserted into thedevice.

The membrane is preferably coated on one or both sides with cells,molecules or other matter, whereby the device provides a controlledenvironment to monitor cell behavior along and/or between themicrochannels via the membrane. One can use any cells from amulticellular organisms in the device. For example, human body comprisesat least 210 known types of cells. A skilled artisan can easilyconstruct useful combinations of the cells in the device. Cell types(e.g., human) which can be used in the devices include, but are notlimited to cells of the integumentary system including but not limitedto Keratinizing epithelial cells, Epidermal keratinocyte(differentiating epidermal cell), Epidermal basal cell (stem cell),Keratinocyte of fingernails and toenails, Nail bed basal cell (stemcell), Medullary hair shaft cell, Cortical hair shaft cell, Cuticularhair shaft cell, Cuticular hair root sheath cell, Hair root sheath cellof Huxley's layer, Hair root sheath cell of Henle's layer, External hairroot sheath cell, Hair matrix cell (stem cell); Wet stratified barrierepithelial cells, such as Surface epithelial cell of stratified squamousepithelium of cornea, tongue, oral cavity, esophagus, anal canal, distalurethra and vagina, basal cell (stem cell) of epithelia of cornea,tongue, oral cavity, esophagus, anal canal, distal urethra and vagina,Urinary epithelium cell (lining urinary bladder and urinary ducts);Exocrine secretory epithelial cells, such as Salivary gland mucous cell(polysaccharide-rich secretion), Salivary gland serous cell(glycoprotein enzyme-rich secretion), Von Ebner's gland cell in tongue(washes taste buds), Mammary gland cell (milk secretion), Lacrimal glandcell (tear secretion), Ceruminous gland cell in ear (wax secretion),Eccrine sweat gland dark cell (glycoprotein secretion), Eccrine sweatgland clear cell (small molecule secretion), Apocrine sweat gland cell(odoriferous secretion, sex-hormone sensitive), Gland of Moll cell ineyelid (specialized sweat gland), Sebaceous gland cell (lipid-rich sebumsecretion), Bowman's gland cell in nose (washes olfactory epithelium),Brunner's gland cell in duodenum (enzymes and alkaline mucus), Seminalvesicle cell (secretes seminal fluid components, including fructose forswimming sperm), Prostate gland cell (secretes seminal fluidcomponents), Bulbourethral gland cell (mucus secretion), Bartholin'sgland cell (vaginal lubricant secretion), Gland of Littre cell (mucussecretion), Uterus endometrium cell (carbohydrate secretion), Isolatedgoblet cell of respiratory and digestive tracts (mucus secretion),Stomach lining mucous cell (mucus secretion), Gastric gland zymogeniccell (pepsinogen secretion), Gastric gland oxyntic cell (hydrochloricacid secretion), Pancreatic acinar cell (bicarbonate and digestiveenzyme secretion), pancreatic endocrine cells, Paneth cell of smallintestine (lysozyme secretion), intestinal epithelial cells, Types I andII pneumocytes of lung (surfactant secretion), and/or Clara cell oflung.

One can also coat the membrane with Hormone secreting cells, such asendocrine cells of the islet of Langerhands of the pancreas, Anteriorpituitary cells, Somatotropes, Lactotropes, Thyrotropes, Gonadotropes,Corticotropes, Intermediate pituitary cell, secretingmelanocyte-stimulating hormone; and Magnocellular neurosecretory cellssecreting oxytocin or vasopressin; Gut and respiratory tract cellssecreting serotonin, endorphin, somatostatin, gastrin, secretin,cholecystokinin, insulin, glucagon, bombesin; Thyroid gland cells suchas thyroid epithelial cell, parafollicular cell, Parathyroid glandcells, Parathyroid chief cell, Oxyphil cell, Adrenal gland cells,chromaffin cells secreting steroid hormones (mineralcorticoids and glucocorticoids), Leydig cell of testes secreting testosterone, Theca internacell of ovarian follicle secreting estrogen, Corpus luteum cell ofruptured ovarian follicle secreting progesterone, Granulosa luteincells, Theca lutein cells, Juxtaglomerular cell (renin secretion),Macula densa cell of kidney, Peripolar cell of kidney, and/or Mesangialcell of kidney.

Additionally or alternatively, one can treat at least one side of themembrane with Metabolism and storage cells such as Hepatocyte (livercell), White fat cell, Brown fat cell, Liver lipocyte. One can also useBarrier function cells (Lung, Gut, Exocrine Glands and Urogenital Tract)or Kidney cells such as Kidney glomerulus parietal cell, Kidneyglomerulus podocyte, Kidney proximal tubule brush border cell, Loop ofHenle thin segment cell, Kidney distal tubule cell, and/or Kidneycollecting duct cell.

Other cells that can be used in the device include Type I pneumocyte(lining air space of lung), Pancreatic duct cell (centroacinar cell),Nonstriated duct cell (of sweat gland, salivary gland, mammary gland,etc.), principal cell, Intercalated cell, Duct cell (of seminal vesicle,prostate gland, etc.), Intestinal brush border cell (with microvilli),Exocrine gland striated duct cell, Gall bladder epithelial cell,Ductulus efferens nonciliated cell, Epididymal principal cell, and/orEpididymal basal cell.

One can also use Epithelial cells lining closed internal body cavitiessuch as Blood vessel and lymphatic vascular endothelial fenestratedcell, Blood vessel and lymphatic vascular endothelial continuous cell,Blood vessel and lymphatic vascular endothelial splenic cell, Synovialcell (lining joint cavities, hyaluronic acid secretion), Serosal cell(lining peritoneal, pleural, and pericardial cavities), Squamous cell(lining perilymphatic space of ear), Squamous cell (lining endolymphaticspace of ear), Columnar cell of endolymphatic sac with microvilli(lining endolymphatic space of ear), Columnar cell of endolymphatic sacwithout microvilli (lining endolymphatic space of ear), Dark cell(lining endolymphatic space of ear), Vestibular membrane cell (liningendolymphatic space of ear), Stria vascularis basal cell (liningendolymphatic space of ear), Stria vascularis marginal cell (liningendolymphatic space of ear), Cell of Claudius (lining endolymphaticspace of ear), Cell of Boettcher (lining endolymphatic space of ear),Choroid plexus cell (cerebrospinal fluid secretion), Pia-arachnoidsquamous cell, Pigmented ciliary epithelium cell of eye, Nonpigmentedciliary epithelium cell of eye, and/or Corneal endothelial cell.

The following cells can be used in the device by adding them to thesurface of the membrane in culture medium. These cells include cellssuch as Ciliated cells with propulsive function such as Respiratorytract ciliated cell, Oviduct ciliated cell (in female), Uterineendometrial ciliated cell (in female), Rete testis ciliated cell (inmale), Ductulus efferens ciliated cell (in male), and/or Ciliatedependymal cell of central nervous system (lining brain cavities).

One can also plate cells that secrete specialized ECMs, such asAmeloblast epithelial cell (tooth enamel secretion), Planum semilunatumepithelial cell of vestibular apparatus of ear (proteoglycan secretion),Organ of Corti interdental epithelial cell (secreting tectorial membranecovering hair cells), Loose connective tissue fibroblasts, Cornealfibroblasts (corneal keratocytes), Tendon fibroblasts, Bone marrowreticular tissue fibroblasts, Other nonepithelial fibroblasts, Pericyte,Nucleus pulposus cell of intervertebral disc, Cementoblast/cementocyte(tooth root bonelike cementum secretion), Odontoblast/odontocyte (toothdentin secretion), Hyaline cartilage chondrocyte, Fibrocartilagechondrocyte, Elastic cartilage chondrocyte, Osteoblast/osteocyte,Osteoprogenitor cell (stem cell of osteoblasts), Hyalocyte of vitreousbody of eye, Stellate cell of perilymphatic space of ear, Hepaticstellate cell (Ito cell), and/or Pancreatic stellate cell.

Additionally or alternatively, contractile cells, such as Skeletalmuscle cells, Red skeletal muscle cell (slow), White skeletal musclecell (fast), Intermediate skeletal muscle cell, nuclear bag cell ofmuscle spindle, nuclear chain cell of muscle spindle, Satellite cell(stem cell), Heart muscle cells, Ordinary heart muscle cell, Nodal heartmuscle cell, Purkinje fiber cell, Smooth muscle cell (various types),Myoepithelial cell of iris, Myoepithelial cell of exocrine glands can beused in the present device.

The following cells can also be used in the present device: Blood andimmune system cells, such as Erythrocyte (red blood cell), Megakaryocyte(platelet precursor), Monocyte, Connective tissue macrophage (varioustypes), Epidermal Langerhans cell, Osteoclast (in bone), Dendritic cell(in lymphoid tissues), Microglial cell (in central nervous system),Neutrophil granulocyte, Eosinophil granulocyte, Basophil granulocyte,Mast cell, Helper T cell, Suppressor T cell, Cytotoxic T cell, NaturalKiller T cell, B cell, Natural killer cell, Reticulocyte, Stem cells andcommitted progenitors for the blood and immune system (various types).One can use these cells as single cell types or in mixtures to determineeffects of the immune cells in the tissue culture system.

One can also treat the membranes with one or more Nervous system cells,Sensory transducer cells such as Auditory inner hair cell of organ ofCorti, Auditory outer hair cell of organ of Corti, Basal cell ofolfactory epithelium (stem cell for olfactory neurons), Cold-sensitiveprimary sensory neurons, Heat-sensitive primary sensory neurons, Merkelcell of epidermis (touch sensor), Olfactory receptor neuron,Pain-sensitive primary sensory neurons (various types); Photoreceptorcells of retina in eye including Photoreceptor rod cells, Photoreceptorblue-sensitive cone cell of eye, Photoreceptor green-sensitive cone cellof eye, Photoreceptor red-sensitive cone cell of eye, Proprioceptiveprimary sensory neurons (various types); Touch-sensitive primary sensoryneurons (various types); Type I carotid body cell (blood pH sensor);Type II carotid body cell (blood pH sensor); Type I hair cell ofvestibular apparatus of ear (acceleration and gravity); Type II haircell of vestibular apparatus of ear (acceleration and gravity); and/orType I taste bud cell.

One can further use Autonomic neuron cells such as Cholinergic neuralcell (various types), Adrenergic neural cell (various types),Peptidergic neural cell (various types) in the present device. Further,sense organ and peripheral neuron supporting cells can also be used.These include, for example, Inner pillar cell of organ of Corti, Outerpillar cell of organ of Corti, Inner phalangeal cell of organ of Corti,Outer phalangeal cell of organ of Corti, Border cell of organ of Corti,Hensen cell of organ of Corti, Vestibular apparatus supporting cell.Type I taste bud supporting cell, Olfactory epithelium supporting cell,Schwann cell, Satellite cell (encapsulating peripheral nerve cellbodies) and/or Enteric glial cell. In some embodiments, one can also usecentral nervous system neurons and glial cells such as Astrocyte(various types), Neuron cells (large variety of types, still poorlyclassified), Oligodendrocyte, and Spindle neuron.

Lens cells such as Anterior lens epithelial cell andCrystallin-containing lens fiber cell can also be used in the presentdevice. Additionally, one can use pigment cells such as melanocytes andretinal pigmented epithelial cells; and germ cells, such asOogonium/Oocyte, Spermatid, Spermatocyte, Spermatogonium cell (stem cellfor spermatocyte), and Spermatozoon.

In some embodiments one can add to the membrane nurse cells Ovarianfollicle cell, Sertoli cell (in testis), Thymus epithelial cell. One canalso use interstitial cells such as interstitial kidney cells.

In an embodiment, one can coat at least one side of the membrane withepithelial cells. Epithelium is a tissue composed of cells that line thecavities and surfaces of structures throughout the body. Many glands arealso formed from epithelial tissue. It lies on top of connective tissue,and the two layers are separated by a basement membrane. In humans,epithelium is classified as a primary body tissue, the other ones beingconnective tissue, muscle tissue and nervous tissue. Epithelium is oftendefined by the expression of the adhesion molecule e-cadherin (asopposed to n-cadherin, which is used by neurons and cells of theconnective tissue).

Functions of epithelial cells include secretion, selective absorption,protection, transcellular transport and detection of sensation and theycommonly as a result present extensive apical-basolateral polarity (e.g.different membrane proteins expressed) and specialization. Examples ofepithelial cells include squamous cells that have the appearance ofthin, flat plates. They fit closely together in tissues; providing asmooth, low-friction surface over which fluids can move easily. Theshape of the nucleus usually corresponds to the cell form and helps toidentify the type of epithelium. Squamous cells tend to havehorizontally flattened, elliptical nuclei because of the thin flattenedform of the cell. Classically, squamous epithelia are found liningsurfaces utilizing simple passive diffusion such as the alveolarepithelium in the lungs. Specialized squamous epithelia also form thelining of cavities such as the blood vessels (endothelium) and heart(mesothelium) and the major cavities found within the body.

Another example of epithelial cells is cuboidal cells. Cuboidal cellsare roughly cuboidal in shape, appearing square in cross section. Eachcell has a spherical nucleus in the centre. Cuboidal epithelium iscommonly found in secretive or absorptive tissue: for example the(secretive) exocrine gland the pancreas and the (absorptive) lining ofthe kidney tubules as well as in the ducts of the glands. They alsoconstitute the germinal epithelium which produces the egg cells in thefemale ovary and the sperm cells in the male testes.

Yet another type of epithelial cells are columnar epithelial cells thatare elongated and column-shaped. Their nuclei are elongated and areusually located near the base of the cells. Columnar epithelium formsthe lining of the stomach and intestines. Some columnar cells arespecialised for sensory reception such as in the nose, ears and thetaste buds of the tongue. Goblet cells (unicellular glands) are foundbetween the columnar epithelial cells of the duodenum. They secretemucus, which acts as a lubricant.

Still another example of the epithelial cells are pseudostratifiedcells. These are simple columnar epithelial cells whose nuclei appear atdifferent heights, giving the misleading (hence “pseudo”) impressionthat the epithelium is stratified when the cells are viewed in crosssection. Pseudostratified epithelium can also possess fine hair-likeextensions of their apical (luminal) membrane called cilia. In thiscase, the epithelium is described as “ciliated” pseudostratifiedepithelium. Cilia are capable of energy dependent pulsatile beating in acertain direction through interaction of cytoskeletal microtubules andconnecting structural proteins and enzymes. The wafting effect producedcauses mucus secreted locally by the goblet cells (to lubricate and totrap pathogens and particles) to flow in that direction (typically outof the body). Ciliated epithelium is found in the airways (nose,bronchi), but is also found in the uterus and Fallopian tubes offemales, where the cilia propel the ovum to the uterus.

Epithelium lines both the outside (skin) and the inside cavities andlumen of bodies. The outermost layer of our skin is composed of deadstratified squamous, keratinised epithelial cells.

Tissues that line the inside of the mouth, the oesophagus and part ofthe rectum are composed of nonkeratinized stratified squamousepithelium. Other surfaces that separate body cavities from the outsideenvironment are lined by simple squamous, columnar, or pseudostratifiedepithelial cells. Other epithelial cells line the insides of the lungs,the gastrointestinal tract, the reproductive and urinary tracts, andmake up the exocrine and endocrine glands. The outer surface of thecornea is covered with fast-growing, easily-regenerated epithelialcells. Endothelium (the inner lining of blood vessels, the heart, andlymphatic vessels) is a specialized form of epithelium. Another type,mesothelium, forms the walls of the pericardium, pleurae, andperitoneum.

Accordingly, one can recreate any of these tissues in the cell culturedevice as described by plating applicable cell types on the porousmembranes and applying applicable vacuum to provide physiological orartificial mechanical force on the cells to mimic physiological forces,such as tension on skin or mechanical strain on lung. In an embodiment,one side of the membrane is coated with epithelial cells and the otherside is coated with endothelial cells.

The endothelium is the thin layer of cells that line the interiorsurface of blood vessels, forming an interface between circulating bloodin the lumen and the rest of the vessel wall. Endothelial cells line theentire circulatory system, from the heart to the smallest capillary.These cells reduce turbulence of the flow of blood allowing the fluid tobe pumped farther. Endothelial tissue is a specialized type ofepithelium tissue (one of the four types of biological tissue inanimals). More specifically, it is simple squamous epithelium.

The foundational model of anatomy makes a distinction betweenendothelial cells and epithelial cells on the basis of which tissuesthey develop from and states that the presence of vimentin rather thankeratin filaments separate these from epithelial cells. Endothelium ofthe interior surfaces of the heart chambers are called endocardium. Bothblood and lymphatic capillaries are composed of a single layer ofendothelial cells called a monolayer. Endothelial cells are involved inmany aspects of vascular biology, including: vasoconstriction andvasodilation, and hence the control of blood pressure; blood clotting(thrombosis & fibrinolysis); atherosclerosis; formation of new bloodvessels (angiogenesis); inflammation and barrier function—theendothelium acts as a selective barrier between the vessel lumen andsurrounding tissue, controlling the passage of materials and the transitof white blood cells into and out of the bloodstream. Excessive orprolonged increases in permeability of the endothelial monolayer, as incases of chronic inflammation, may lead to tissue oedema/swelling. Insome organs, there are highly differentiated endothelial cells toperform specialized ‘filtering’ functions. Examples of such uniqueendothelial structures include the renal glomerulus and the blood-brainbarrier.

In an embodiment, the membrane side that contains culturedendothelialcells can be exposed to various test substances and also white bloodcells or specific immune system cells to study effects of the testagents on the function of the immune system cells at the tissue level.

Details on how the tissue interface device 200 is formed will now bediscussed in accordance with an embodiment. The fabrication of the PDMSmembrane preferably involves parallel processing of multiple parts whichare assembled in stages. FIG. 4A illustrates a perspective view of amaster 600 in accordance with an embodiment which is ultimately used toproduce the porous membrane 208. As shown in FIG. 4A, the master 600 ispreferably formed by patterning a photoresist to the desired shape andsize on a silicon substrate.

It should be noted that the posts 602 may be designed in any desiredarray depending on the intended design of the membrane 208. For example,the posts 602 may be arranged in a circular pattern to correspondinglyform a circular patterned set of pores in the membrane 208. It should benoted that the posts 602 may have any other cross sectional shape otherthan pentagonal to make the corresponding pores in the membrane, asdiscussed above. It should also be noted that the master 600 may containdifferent height ridges to create non planar membranes.

Thereafter, as shown in FIG. 4B, the master 600 is preferablyspin-coated with PDMS to form a spin coated layer 604. Thereafter, thespin-coated layer 604 is cured for a set time and temperature (e.g. 110°C. at 15 minutes) and peeled off the master 600 to produce a thin PDMSmembrane 604 having the array of pentagonal through-holes 606, as shownin FIG. 4C. The example shown depicts fabrication of a 10 μm-thick PDMSmembrane, although other thickness values are contemplated.

Although other materials may be used, PDMS has useful properties inbiology in that it is a moderately stiff elastomer (1 MPa) which isnon-toxic and is optically transparent to 300 nm. PDMS is intrinsicallyvery hydrophobic, but can be converted to hydrophilic form by treatmentwith plasma. The membrane 604 may be engineered for a variety ofpurposes, some discussed above. For example, the pores 606 on themembrane 604 may be coated or filled with ECM molecules or gels, such asMATRIGEL, laminin, collagen, fibronectin, fibrin, elastin, etc., whichare known to those skilled in the art. The tissue-tissue interface maybe coated by culturing different types of cells on each side of themembrane 604, as shown in FIG. 4D. In particular, as shown in FIG. 4D,one type of cells 608 are coated on one side of the membrane 604 whereasanother type of cells 610 are coated on the opposing side of themembrane 604.

FIGS. 5A and 5B illustrate the process how the first outer body portion202, a second outer body portion 204 are formed in accordance with anembodiment. The first and second outer body portions 202, 204 arepreferably formed using soft lithography techniques, although othertechniques well known in the art are contemplated. In an embodiment, aphotoresist (not shown) is formed on a substrate in which thephotoresist has positive relief features which mirror the desiredbranching configuration in the first outer body portion. Similarly, asecond photoresist (not shown) is formed on another substrate in whichthe second photoresist has corresponding positive relief features whichmirror the branching configuration in the second outer body portion 204.The microchannels along with the communicating ports and port aperturesare preferably generated by preferably casting PDMS or other appropriatematerial onto each master. Once the first and second outer body portions202, 204 are formed, through-holes which serve as the port apertures aremade through the PDMS slab preferably using an aperture formingmechanism or stamp.

As shown in FIG. 5C, the already formed PDMS membrane 208 is thensandwiched between the first outer body portion 202 and the second outerbody portion 204, whereby the microchannel walls 234, 244 as well as theoutside walls 238, 248 are aligned using appropriate manufacturingequipment and techniques. Thereafter, the microchannel walls 234, 244and outside walls are preferably bonded to the membrane 208 using anappropriate adhesive or epoxy. Additionally, the remaining portions ofthe outer body portions 202, 204 are permanently bonded to one anotherusing an appropriate adhesive or epoxy to form the overall device.

Subsequently, as shown in FIG. 5D, a PDMS etching solution is introducedinto the operating channels to etch away the PDMS membrane segments inthe operating channels. This results in resulting in the generation ofthe two side operating channels 252 being free from the membrane,although the membrane is maintained in the central microchannel, asshown in FIG. 5E. The above is preferably formed using soft lithographytechniques, the details of which are described in “Soft Lithography inBiology and Biochemistry,” by Whitesides, et al., published AnnualReview, Biomed Engineering, 3.335-3.373 (2001), as well as “AnUltra-Thin PDMS Membrane As A Bio/Micro-Nano Interface: Fabrication AndCharacterization”, by Thangawng et al., Biomed Microdevices, vol. 9,num. 4, 2007, p. 587-95, both of which are hereby incorporated byreference.

FIG. 6 illustrates a schematic of a system having multiple tissueinterface devices in accordance with an embodiment. In particular, asshown in FIG. 6, the system 700 includes one or more CPUs 702 coupled toone or more fluid sources 704 and pressure sources (not shown), wherebythe preceding are coupled to three shown tissue interface devices 706A,706B, and 706C. It should be noted that although three devices 706 areshown in this embodiment, fewer or greater than three devices 706 arecontemplated. In the system 700, two of the three devices (i.e. 706A and706B) are connected in parallel with respect to the fluid source 704 anddevices 706A and 706C are connected in serial fashion with respect tothe fluid source 704. It should be noted that the shown configuration isonly one example and any other types of connection patterns may beutilized depending on the application.

In the example shown, fluid from the fluid source 704 is provideddirectly to the fluid inlets of devices 706A and 706B. As the fluidpasses through device 706A, it is output directly into the fluid inletport of devices 706B and 706C. Additionally, the fluid outlet fromdevice 706B is combined with the output from device 706A into device706C. With multiple devices operating, it is possible to monitor, usingsensor data, how the cells in the fluid or membrane behave after thefluid has been passed through another controlled environment. Thissystem thus allows multiple independent “stages” to be set up, wherecell behavior in each stage may be monitored under simulatedphysiological conditions and controlled using the devices 706. One ormore devices are connected serially may provide use in studying chemicalcommunication between cells. For example, one cell type may secreteprotein A in response to being exposed to a particular fluid, wherebythe fluid, containing the secreted protein A, exits one device and thenis exposed to another cell type specifically patterned in anotherdevice, whereby the interaction of the fluid with protein A with theother cells in the other device can be monitored (e.g. paracrinesignaling). For the parallel configuration, one or more devicesconnected in parallel may be advantageous in increasing the efficiencyof analyzing cell behavior across multiple devices at once instead ofanalyzing the cell behavior through individual devices separately.

FIG. 7A illustrates a perspective view of an organ mimic device inaccordance with an embodiment that contains three parallel microchannelsseparated by two porous membranes. As shown in FIG. 7A, the organ mimicdevice 800 includes operating microchannels 802 and an overall centralmicrochannel 804 positioned between the operating microchannels 802. Theoverall central microchannel 804 includes multiple membranes 806A, 806Bpositioned along respective parallel x-y planes which separate themicrochannel 804 into three distinct central microchannels 804A, 804Band 804C. The membranes 806A and 806B may be porous, elastic, or acombination thereof. Positive and/or negative pressurized media may beapplied via operating channels 802 to create a pressure differential tothereby cause the membranes 806A, 806B to expand and contract alongtheir respective planes in parallel.

FIG. 7B illustrates a perspective view of an organ mimic device inaccordance with an embodiment. As shown in FIG. 7B, the tissue interfacedevice 900 includes operating microchannels 902A, 902B and a centralmicrochannel 904 positioned between the microchannels 902. The centralmicrochannel 904 includes multiple membranes 906A, 906B positioned alongrespective parallel x-y planes. Additionally, a wall 910 separates thecentral microchannel into two distinct central microchannels, havingrespective sections, whereby the wall 910 along with membranes 904A and904B define microchannels 904A, 904B, 904C, and 904D. The membranes 906Aand 906B at least partially porous, elastic or a combination thereof.

The device in FIG. 7B differs from that in FIG. 7A in that the operatingmicrochannels 902A and 902B are separated by a wall 908, wherebyseparate pressures applied to the microchannels 902A and 902B causetheir respective membranes 904A and 904B to expand or contract. Inparticular, a positive and/or negative pressure may be applied viaoperating microchannels 902A to cause the membrane 906A to expand andcontract along its plane while a different positive and/or negativepressure is applied via operating microchannels 902B to cause themembrane 906B to expand and contract along its plane at a differentfrequency and/or magnitude. Of course, one set of operatingmicrochannels may experience the pressure while the other set does notexperience a pressure, thereby only causing one membrane to actuate. Itshould be noted that although two membranes are shown in the devices 800and 900, more than two membranes are contemplated and can be configuredin the devices.

In an example, shown in FIG. 7C, the device containing three channelsdescribed in FIG. 7A has two membranes 806A and 806B which are coated todetermine cell behavior of a vascularized tumor. In particular, membrane806A is coated with a lymphatic endothelium on its upper surface 805Aand with stromal cells on its lower surface, and stromal cells are alsocoated on the upper surface of the second porous membrane 805B and avascular endothelium on its bottom surface 805C. Tumor cells are placedin the central microchannel surrounded on top and bottom by layers ofstromal cells on the surfaces of the upper and lower membranes insection 804B. Fluids such as cell culture medium or blood enters thevascular channel in section 804 C (you are missing a label 804C in thediagram). Fluid such as cell culture medium or lymph enters thelymphatic channel in section 804A. This configuration of the device 800allows researchers to mimic and study tumor growth and invasion intoblood and lymphatic vessels during cancer metastasis. In the example,one or more of the membranes 806A, 806B may expand/contract in responseto pressure through the operating microchannels. Additionally oralternatively, the membranes may not actuate, but may be porous or havegrooves to allow cells to pass through the membranes.

The unique capabilities of the present device have been monitored inexperiments that address acute toxicity and extrapulmonary translocationof engineered nanomaterials induced by physiological mechanical forces.The device has been used to model pulmonary inflammation in which it canprecisely recreate and directly visualize the complex interplay ofpulmonary tissues with cytokines and blood-borne immune cells thattransmigrate across the alveolar-capillary barrier. Using this model,the device reveals significant inflammatory responses of the mimickedlung to nanomaterials. Finally, the device is used to simulate pulmonaryinfection with bacteria and its clearance by neutrophil recruitment andphagocytosis.

The device has been used in experiments which have led to the discoverythat physiological mechanical forces can induce or exacerbate toxicityof engineered nanomaterials in the lung and may facilitate theirtranslocation into the systemic circulation. Furthermore, in vitromodels that simulate lung inflammation have been developed that enabledirect observation of the adhesion of circulating blood-borne immunecells to inflamed endothelia and their transmigration across thealveolar-capillary barrier. Based on this model, significantproinflammatory activities of engineered nanoparticles have beenrevealed. Based on this evidence, a model of pulmonary infection can beestablished and re-creation may be done of the innate immune response ofthe lung to bacteria mediated by neutrophil infiltration into thealveoli and bacterial phagocytosis.

The present device was utilized in several experiments, whereby thedevice was used to mimic the living lung. The observations and findingswith the present device are described hereafter. During normalinspiration of a real lung, the thoracic cavity enlarges due to thecontraction of the diaphragm and expansion of the rib-cage and, as aresult, the intrapleural pressure outside the alveoli decreases. Theincreased pressure difference across the alveolar wall causes thealveoli to expand and forces air into the lungs, resulting in stretchingof the alveolar epithelium and endothelium in the surroundingcapillaries. Alveolar epithelial cells are co-cultured with pulmonarymicrovascular endothelial cells on a thin porous membrane to produce twoopposing tissue layers that mimic the interface between the alveolarepithelium and pulmonary endothelium. The compartmentalized microchannelconfiguration makes it readily possible to manipulate fluidicenvironment of the epithelium and endothelium independently, and toapply physiological mechanical strain.

In the experiment, co-culture of alveolar epithelial cells and primarypulmonary microvascular endothelial cells of human origin was developedover two weeks without loss of viability. The microfluidic cultureresulted in the production of tight alveolar-capillary barriers withstructural integrity as evidenced by typical junctional complexespresent in both epithelial and endothelial layers. The microfluidicdevice was integrated with computer-controlled vacuum to enable cyclicmembrane/cell stretching at varying frequencies and levels of strain ina programmable manner. It was observed that applied vacuum generatedunidirectional tension which is uniform across the wide centralmicrochannel. Concurrently, it was discovered that this tension wasperceived by adherent cells and caused them to stretch and increasetheir projected surface area. Also effective application of mechanicalstrain to cells was confirmed by showing stretch-induced alignment andtransient calcium responses of endothelial cells.

Based on the unique capabilities afforded by on-chip production ofpulmonary tissues and faithful recapitulation of their nativemicroenvironment, the device was used to assess the potential adverseeffects of nanomaterials. Despite the widespread use of engineerednanomaterials, much remains to be learned about their risks to healthand environment. Existing toxicology methods rely on oversimplified invitro models or lengthy, expensive animal testing that often poseschallenges to mechanistic studies at the cellular level. To bridge thegap between cell culture studies and animal models, the device was usedto permit a more realistic, accurate evaluation of nanomaterial toxicityin a tightly controlled biomimetic microenvironment.

In the experiment, the alveolar epithelial tissues prepared in thedevice were exposed to various nanomaterials and oxidative stress wasexamined by measuring intracellular production of reactive oxygenspecies (ROS) using microfluorimetry. Through the testing of colloidalsilica nanoparticles and quantum dots, it was discovered thatphysiological mechanical strain can dramatically increasenanoparticle-generated oxidative stress and induce early toxic responsesin the pulmonary epithelium. For example, when the cells were exposed to12 nm silica nanoparticles in combination with a cyclic stretch of 10%strain at 0.2 Hz which simulates normal respiration, ROS productionincreased by more than five times after two hours, whereas nanoparticlesor mechanical strain alone did not cause any measurable responses overthe duration of the experiments (see FIG. 8). The response of cellstreated with carboxylated quantum dots showed similar trends (see FIG.9). It was noted that similar levels of ROS increase were achieved after24 hour-long exposures to silica nanoparticles alone, as shown in FIG.9.

It was also found that cyclic strain alone did not have any significantimpact regardless of its duration, as shown in FIG. 9. Taken together,these observations suggest that physiological forces act in synergy withnanoparticles to exert early toxic effects or aggravate nanoparticletoxicity in the lung. This stretch-induced ROS response to nanomaterialsdepended on the level of strain and induced apoptosis of the epithelialcells as detected by caspase activity. When treated with a clinicallyused free radical scavenger, N-acetylcysteine (NAC) during nanoparticleexposure, the cells were completely rescued from oxidative stresspresumably due to the antioxidant activity of NAC leading to increasedintracellular glutathione. It was also observed that oxidative stressgenerated by the combined effect of nanomaterials and strain variedsignificantly with the type of nanomaterials. For example, exposures to50 nm superparamagnetic iron nanoparticles under the same conditionsonly resulted in a transient increase in oxidative stress. This uniqueROS response was not observed in the testing of other nanomaterialsincluding single walled carbon nanotubes, gold nanoparticles,polystyrene nanoparticles, and quantum dots coated with polyethyleneglycol, as shown below in Table 1.

TABLE 1 ROS ROS response response (0% (10% Nanomaterials Surface coatingSize strain) strain) Polystyrene Carboxyl groups 500 nm No Nonanoparticles Carboxyl groups 200 nm No No Amine groups 200 nm No NoCarboxyl groups 100 nm No No Carboxyl groups 20 nm No No Quantum dotsCarboxyl groups 16 nm No Yes polyethylene 13 nm No No glycol Silica N/A12 nm No Yes nanoparticles Magnetic iron Carboxyl groups 50 nm No Yesnanoparticles Gold N/A 3 nm No No nanoparticles

To understand the influence of physiological forces ontissue-nanomaterial interactions, confocal microscopy was used toanalyze internalization of 100 nm fluorescent nanoparticles into theepithelial cells after 1 hour of exposure. However, the number ofparticles or their aggregates detected in intracellular compartments wasmuch greater in the presence of mechanical strain, and over 80% of thecells were found to have taken up the nanoparticles, whereas the extentof nanoparticle uptake was considerably smaller in the absence ofstrain. These results indicate that physiological mechanical forces mayfacilitate cellular uptake of nanomaterials, allowing them to interactwith subcellular components and thereby rendering them potentially moreharmful.

Moreover, the device provides an opportunity to investigateextrapulmonary translocation of nanomaterials from the alveolar space tothe microvasculature. Increasing in vivo evidence suggests thatnanomaterials in the alveoli have the capacity to cross thealveolar-capillary barrier and enter the pulmonary circulation,potentially impacting other organs. To investigate this situation, 20 nmfluorescent nanoparticles were introduced on the epithelial side andnanoparticle translocation was monitored by counting the number ofparticles carried out of the lower vascular channel by continuous fluidflow. This model revealed a marked increase in the rate of nanoparticlemigration into the vascular compartment under physiological conditionswith 10% cyclic strain, as compared to transport across a relaxed,static tissue barrier. These findings provide in vitro evidence that theinherent mechanical activity of the living lung may allow nanomaterialsto translocate from the alveolar space into the bloodstream. The datafrom the experiment also supports the systematic distribution andaccumulation of inhaled nanomaterials observed in animal studies and maypotentially contribute to delineating the mechanism of this process, aswell as providing a surrogate model system for studying this response.

To further demonstrate the device's capabilities to reconstitute theintegrated organ-level responses in the lung, a more sophisticated modelwas developed that incorporated circulating blood-borne immune cells andreproduced the key steps of lung inflammation. Generally, inflammatoryresponses in the lung involve a highly coordinated multistep cascade ofepithelial production and release of early response cytokines,activation of vascular endothelium through upregulation of leukocyteadhesion molecules and subsequent leukocyte infiltration from thepulmonary microcirculation into the alveolar space. To simulate thisprocess, the apical surface of the alveolar epithelium was firststimulated with tumor necrosis factor-α (TNF-α), which is a potentpro-inflammatory mediator, and endothelial activation was examined bymeasuring the expression of intercellular adhesion molecule-1 (ICAM-1).In response to TNF-α stimulation of the alveolar tissue for 5 hours, theendothelial cells on the opposite side of the membrane dramaticallyincreased their surface expression of ICAM-1. Furthermore, the activatedendothelium supported capture and firm adhesion of human neutrophilsflowing in the vascular microchannel, which did not adhere in theabsence of cytokine exposure. Treatment of the epithelial cells with lowdoses of TNF-α resulted in weak activation of the endothelium, whichcaused captured neutrophils to roll continuously in the direction offlow without being arrested. Direct microscopic visualization revealedthat adherent neutrophils became flattened and crawled from a site offirm adhesion to distant locations where they extravasated through theendothelium and transmigrated across the alveolar-capillary barrierthrough the membrane pores over the period of several minutes. Thetransmigrated neutrophils then emigrated onto the apical surface of thealveolar epithelium preferentially through paracellular junctions andwere retained on the epithelial layer in spite of fluid flow and cyclicstretching. These sequential events successfully replicate the entireprocess of neutrophil recruitment from the microvasculature to thealveolar compartment, which is a hallmark of lung inflammation.

Using the device, proinflammatory effects of colloidal silicananoparticles on the lung were investigated. Upon the alveolarepithelial cells being exposed to 12 nm silica nanoparticles for 5hours, the microvascular endothelium became activated and exhibited highlevels of ICAM-1 expression. It was noted that application of 10% cyclicstrain along with nanoparticles synergistically upregulated endothelialexpression of ICAM-1. Human neutrophils circulating in the vascularchannel were seen to firmly adhere to the inflamed endothelium, totransmigrate across the tissue barrier, and to accumulate on theepithelial surface. These observations evidence significantproinflammatory activities of these silica nanoparticles, which maybecome more pronounced due to physiological forces that provoke acuteinflammation in the lung.

In an experiment, the present device was configured to mimic the innateimmune response to pulmonary infection of bacterial origin. To imitatethe lung afflicted with bacterial infection, alveolar epithelial cellswere apically stimulated with Escherichia coli (E. coli) constitutivelyexpressing green fluorescent protein (GFP) for 5 hours. When humanneutrophils were subsequently allowed to flow in the vascularmicrochannel, they attached to the endothelial cells and underwentdiapedesis across the tissue layers, indicating that bacterialstimulation of the epithelium gave rise to endothelial activation. Uponreaching the epithelial surface, the neutrophils showed directionalmovement towards GFP-labeled bacteria and engulfed them as illustratedby detection of phagocytosed bacteria with fluorescently labeled movingneutrophils. It was also observed that neutrophils are capable ofingesting more than one bacterium over short periods of time and thattheir phagocytic activity continued until a majority of the bacteriawere cleared from the observation area. These results clearlydemonstrate the ability of this model to recreate the complete processof the integrated immune response to microbial infection within a 3Dphysiological organ context in vitro.

While embodiments and applications have been shown and described, itwould be apparent to those skilled in the art having the benefit of thisdisclosure that many more modifications than mentioned above arepossible without departing from the inventive concepts disclosed herein.The embodiment(s), therefore, are not to be restricted except in thespirit of the appended claims.

The present inventive subject matter can be defined in any of thefollowing alphabetized paragraphs:

[A] An organomimetic device comprising:

a body having a central microchannel therein; and

an at least partially porous membrane positioned within the centralmicrochannel and along a plane, the membrane configured to separate thecentral microchannel to form a first central microchannel and a secondcentral microchannel, wherein a first fluid is applied through the firstcentral microchannel and a second fluid is applied through the secondcentral microchannel, the membrane coated with at least one attachmentmolecule that supports adhesion of a plurality of living cells.

[B] The device of [A] wherein the porous membrane is at least partiallyflexible, the device further comprising:

a first chamber wall of the body positioned adjacent to the first andsecond central microchannels, wherein the membrane is mounted to thefirst chamber wall; and

a first operating channel adjacent to the first and second centralmicrochannels on an opposing side of the first chamber wall, wherein apressure differential applied between the first operating channel andthe central microchannels causes the first chamber wall to flex in afirst desired direction to expand or contract along the plane within thefirst and second central microchannels.

[C] The device of [A] or [B] further comprising:

a second chamber wall of the body positioned adjacent to the first andsecond central microchannels, wherein an opposing end of the membrane ismounted to the second chamber wall; and

a second operating channel positioned adjacent to the centralmicrochannel on an opposing side of the second chamber wall, wherein thepressure differential between to the second operating channel and thecentral microchannels causes the second chamber wall to flex in a seconddesired direction to expand or contract along the plane within the firstand second central microchannels.

[D] The device of any or all of the above paragraphs wherein at leastone pore aperture in the membrane is between 0.5 and 20 microns along awidth dimension.

[E] The device of any or all of the above paragraphs wherein themembrane further comprises a first membrane and a second membranepositioned within the central microchannel, wherein the second membraneis oriented parallel to the first membrane to form a third centralmicrochannel therebetween.

[F] The device of any or all of the above paragraphs wherein themembrane comprises PDMS,

[G] The device of any or all of the above paragraphs wherein themembrane is coated with one or more cell layers, wherein the one or morecell layers are applied to a surface of the membrane.

[H] The device of any or all of the above paragraphs wherein one or bothsides of the membrane are coated with one or more cell layers, whereinthe one or more cell layers comprise cells selected from the groupconsisting of metazoan, mammalian, and human cells.

[I] The device of any or all of the above paragraphs, wherein the cellsare selected from the group consisting of epithelial, endothelial,mesenchymal, muscle, immune, neural, and hemapoietic cells.

[J] The device of any or all of the above paragraphs wherein one side ofthe membrane is coated with epithelial cells and the other side of themembrane is coated with endothelial cells.

[K] The device of any or all of the above paragraphs wherein the body ofthe device and the membrane are made of a biocompatible or biodegradablematerial.

[L] The device of any or all of the above paragraphs wherein the deviceis further implanted to a living organism.

[M] The device of any or all of the above paragraphs wherein the livingorganism is a human.

[N] The device of any or all of the above paragraphs wherein themembrane is coated with the one or more cell layers in vitro.

[O] The device of any or all of the above paragraphs, wherein the atleast one membrane is coated with the one or more cell layers in vivo.

[P] The device of any or all of the above paragraphs, wherein themembrane is coated with a biocompatible agent which facilitatesattachment of the at least one cell layer onto the membrane.

[Q] The device of any or all of the above paragraphs wherein thebiocompatible agent is extracellular matrix comprising collagen,fibronectin and/or laminin.

[R] The device of any or all of the above paragraphs wherein thebiocompatible material is selected from the group consisting ofcollagen, laminin, proteoglycan, vitronectin, fibronectin, poly-D-lysineand polysaccharide.

[S] The device of any or all of the above paragraphs wherein the firstfluid contains white blood cells.

[T] A method comprising:

selecting a organomimetic device having a body, the body including an atleast partially porous membrane positioned along a plane within acentral microchannel to partition the central microchannel into a firstcentral microchannel and a second central microchannel, the membranecoated with at least one attachment molecule that supports adhesion of aplurality of living cells;

applying a first fluid through the first central microchannel;

applying a second fluid through the second central microchannel; and

monitoring behavior of cells with respect to the membrane between thefirst and second central micro channels.

[U] The method of any or all of the above paragraphs wherein themembrane is at least partially elastic and the body includes at leastone operating channel positioned adjacent to the first and secondcentral microchannels, the method further comprising:

adjusting a pressure differential between the central microchannels andthe at least one operating channels, wherein the membrane stretchesalong the plane in response to the pressure differential.

[V] The method of any or all of the above paragraphs wherein theadjusting of the pressure differential further comprises:

increasing the pressure differential such that one or more sides of themembrane move in desired directions along the plane; and

decreasing the pressure differential such that the one or more sides ofthe membrane move in an opposite direction along the plane.

[W] The method of any or all of the above paragraphs wherein at leastone pore aperture in the membrane is between 0.5 and 20 microns along awidth dimension.

[X] The method of any or all of the above paragraphs further comprisingtreating the membrane with one or more cell layers, wherein the one ormore cell layers are applied to a surface of the membrane.

[Y] The method of any or all of the above paragraphs further comprisingapplying one or more cell layers onto one or both sides of the membrane,wherein the one or more cell layers comprise cells selected from thegroup consisting of metazoan, mammalian, and human cells.

[Z] The method of any or all of the above paragraphs wherein the cellsare selected from the group consisting of epithelial, endothelial,mesenchymal, muscle, immune, neural, and hemapoietic cells.

[AA] The method of any or all of the above paragraphs wherein one sideof the membrane is coated with epithelial cells and the other side ofthe membrane is coated with endothelial cells.

[BB] The method of any or all of the above paragraphs wherein the bodyof the device and the membrane are made of a biocompatible orbiodegradable material.

[CC] The method of any or all of the above paragraphs wherein the deviceis further implanted to a living organism.

[DD] The method of any or all of the above paragraphs wherein the livingorganism is a human.

[EE] The method of any or all of the above paragraphs wherein themembrane is coated with the one or more cell layers in vitro.

[FF] The method of any or all of the above paragraphs wherein the atleast one membrane is coated with the one or more cell layers in vivo.

[GG] The method of any or all of the above paragraphs wherein themembrane is coated with a biocompatible agent which facilitatesattachment of the at least one cell layer onto the membrane.

[HH] The method of any or all of the above paragraphs wherein thebiocompatible agent is extracellular matrix comprising collagen,fibronectin and/or laminin.

[II] The method of any or all of the above paragraphs wherein thebiocompatible material is selected from the group consisting ofcollagen, laminin, proteoglycan, vitronectin, fibronectin, poly-D-lysineand polysaccharide.

[JJ] The method of any or all of the above paragraphs wherein the firstfluid contains white blood cells.

[KK] A method for determining an effect of at least one agent in atissue system with physiological or pathological mechanical force, themethod comprising:

selecting a device having a body, the body including an at leastpartially porous membrane positioned along a plane within a centralmicrochannel to partition the central microchannel into a first centralmicrochannel and a second central microchannel;

contacting the membrane with at least one layer of cells on a first sideof the membrane and at least one layer of cells on a second side of theporous membrane thereby forming a tissue structure comprising at leasttwo different types of cells;

contacting the tissue structure comprising at least two different typesof cells with the at least one agent in an applicable cell culturemedium;

-   -   applying uniform or non-uniform force on the cells for a time        period; and    -   measuring a response of the cells in the tissue structure        comprising at least two different types of cells to determine        the effect of the at least one agent on the cells.

[LL] The method of any or all of the above paragraphs wherein theapplicable cell culture medium is supplemented with white blood cells.

[MM] The method of any or all of the above paragraphs wherein theuniform or non-uniform force is applied using vacuum.

[NN] The method of any or all of the above paragraphs wherein the tissuestructure comprising at least two different types of cells comprisesalveolar epithelial cells on the first side of the porous membrane andpulmonary microvascular cells on the second side of the porous membrane.

[OO] The method of any or all of the above paragraphs wherein the agentis selected from the group consisting of nanoparticles, environmentaltoxins or pollutant, cigarette smoke, chemicals or particles used incosmetic products, drugs or drug candidates, aerosols, naturallyoccurring particles including pollen, chemical weapons, single ordouble-stranded nucleic acids, viruses, bacteria and unicellularorganisms.

[PP] The method of any or all of the above paragraphs wherein themeasuring the response is performed by measuring expression of reactiveoxygen species.

[QQ] The method of any or all of the above paragraphs wherein themeasuring the response is performed using tissue staining.

[RR] The method of any or all of the above paragraphs further comprisingprior to measuring the effect of the agent, taking a biopsy of themembrane comprising tissue structure comprising at least two differenttypes of cells, wherein the biopsy is stained.

[SS] The method of any or all of the above paragraphs wherein themeasuring the response is performed from a sample of the cell culturemedium in contact wherein the measuring the response is performed from asample of the cell culture medium in contact with the first or thesecond or both sides of the membrane form tissue structure comprising atleast two different types of cells. with the first or the second or bothsides of the membrane comprising tissue structure comprising at leasttwo different types of cells.

[TT] The method of any or all of the above paragraphs further comprisingcomparing the effect of the agent to another agent or a control withoutthe agent in a similar parallel device system.

[UU] The method of any or all of the above paragraphs further comprisinga step of contacting the membrane with at least two agents, wherein thefirst agent is contacted first to cause an effect on the tissuestructure comprising at least two different types of cells and the atleast second agent in contacted after a time period to test the effectof the second agent on the tissue structure comprising at least twodifferent types of cells affected with the first agent.

[VV] An organomimetic device comprising:

a body having a central microchannel; and

a plurality of membranes positioned along parallel planes in the centralmicrochannel, wherein at least one of the plurality of membranes is atleast partially porous, the plurality of membranes configured topartition the central microchannel into a plurality of centralmicrochannels.

1. (canceled)
 2. A device for monitoring a biological function,comprising: a body having a first microchannel and a secondmicrochannel; a membrane located at an interface region between thefirst microchannel and the second microchannel, the membrane including afirst side facing toward the first microchannel and a second side facingtoward the second microchannel, the first side having cells of a firsttype adhered thereto; and a sensor coupled to the membrane.
 3. Thedevice of claim 2, wherein the sensor enables a measurement of anelectrical characteristic across the membrane.
 4. The device of claim 3,wherein the electrical characteristic is a potential difference acrossthe membrane.
 5. The device of claim 3, wherein the electricalcharacteristic is a short-circuit current condition across the membrane.6. The device of claim 3, wherein the electrical characteristic is aresistance across the membrane.
 7. The device of claim 2, wherein thesensor enables a measurement of a characteristic that confirms theformation of an organized barrier of the first type of cells.
 8. Thedevice of claim 2, wherein the sensor enables a measurement of acharacteristic that confirms an ion transport function across themembrane.
 9. The device of claim 2, wherein the sensor enables ameasurement of a characteristic that confirms a fluid transport functionacross the membrane.
 10. The device of claim 2, wherein the sensorincludes one or more microelectrodes that enable a measurement of anelectrical characteristic across the membrane.
 11. The device of claim10, wherein the electrical characteristic confirms a formation of anorganized barrier of the first type of cells.
 12. The device of claim10, wherein the electrical characteristic confirms an ion transportfunction across the membrane.
 13. The device of claim 10, wherein theelectrical characteristic confirms a fluid transport function across themembrane.
 14. The device of claim 2, wherein the membrane iscontrollably stretchable in a desired direction while a first fluid ispresent in the first microchannel and a second fluid is present in thesecond microchannel.
 15. The device of claim 2, wherein the sensorenables a measurement of an electrical resistance across the membrane tomonitor changes in the vascular permeability during inflammatoryresponses of the first type of cells.
 16. The device of claim 2, whereinthe sensor enables a measurement of a short-circuit current conditionacross the membrane to monitor changes in the vascular permeabilityduring inflammatory responses of the first type of cells.
 17. The deviceof claim 2, wherein the sensor enables a measurement of a characteristicfor monitoring transmigration of nanomaterials through the first type ofcells on the membrane.
 18. The device of claim 2, wherein the sensorenables a measurement of a characteristic for monitoringnanomaterial-induced changes in a barrier function associated with thefirst type of cells on the membrane.
 19. The device of claim 2, whereinthe membrane is a made of a material having a plurality of pores orapertures, thereby permitting the migration of at least one of cells,particulates, chemicals, molecules, fluids, liquids, and gases from thefirst side of the membrane to the second side of the membrane.
 20. Thedevice of claim 2, wherein the membrane is made of more than onematerial.
 21. The device of claim 20, wherein the membrane includes acoating.
 22. The device of claim 21, wherein the coating includes ametal.
 23. The device of claim 20, wherein the membrane includes fibers.24. A device for monitoring a biological function, comprising: a bodyhaving a first microchannel and a second microchannel; a membranelocated at an interface region between the first microchannel and thesecond microchannel, the membrane including a first side facing towardthe first microchannel and a second side facing toward the secondmicrochannel, the first side having cells of a first type adheredthereto; and at least one sensor that enables a measurement of anelectrical characteristic across the membrane with the first type ofcells adhered thereto.
 25. The device of claim 24, wherein theelectrical characteristic is a potential difference across the membrane.26. The device of claim 24, wherein the electrical characteristic is ashort-circuit current condition across the membrane.
 27. The device ofclaim 24, wherein the electrical characteristic is a resistance acrossthe membrane.
 28. The device of claim 24, wherein the membrane is madeof more than one material and the sensor is coupled to the membrane. 29.The device of claim 28, wherein the membrane includes a coating.
 30. Thedevice of claim 29, wherein the coating includes a metal.
 31. A systemfor monitoring a biological function, comprising: a device having afirst microchannel, a second microchannel, and a membrane located at aninterface region between the first microchannel and the secondmicrochannel, the membrane including a first side facing toward thefirst microchannel and a second side facing toward the secondmicrochannel, the first side having cells of a first type adheredthereto, the device further including at least one sensor integratedwithin the device; and a processor coupled to the sensor for receivingdata from the sensor.
 32. The system of claim 31, further including adisplay for displaying the data.
 33. The system of claim 31, wherein thedata provides information on an operational condition of the device. 34.The system of claim 31, wherein the data provides information on abehavior of the first type of cells on a real-time basis.
 35. The systemof claim 31, wherein the at least one sensor includes one or moremicroelectrodes that enable a measurement of an electricalcharacteristic across the membrane.
 36. The system of claim 31, whereinthe membrane is controllably stretchable in a desired direction while afirst fluid is present in the first microchannel and a second fluid ispresent in the second microchannel.
 37. The system of claim 31, whereinthe membrane is a made of a material having a plurality of pores orapertures, thereby permitting the migration of at least one of cells,particulates, chemicals, molecules, fluids, liquids, and gases from thefirst side of the membrane to the second side of the membrane.
 38. Thesystem of claim 31, wherein the membrane is made of more than onematerial and the at least one sensor is coupled to the membrane.
 39. Thesystem of claim 38, wherein the membrane includes a coating.
 40. Thesystem of claim 39, wherein the coating includes a metal.
 41. The systemof claim 38, wherein the membrane includes fibers.
 42. The system ofclaim 31, wherein the membrane is at least partially opticallytransparent.
 43. A method of monitoring a biological function in adevice having a membrane located on an interface region between a firstmicrochannel and a second microchannel, a first side of the membranefacing the first microchannel and having a first type of cells adheredthereto, a second side of the membrane facing the second microchannel,the method comprising: moving a first fluid through at least one of thefirst microchannel and the second microchannel; and measuring anelectrical characteristic across the membrane with the first type ofcells adhered thereto.
 44. The method of claim 43, further includingcontrollably stretching the membrane in a first direction so as to applya force to the first type of cells adhered to the first side of themembrane.
 45. The method of claim 43, wherein the membrane includes atleast one sensor coupled thereto, the sensor for measuring theelectrical characteristic.
 46. The method of claim 45, wherein the atleast one sensor includes one or more microelectrodes.
 47. The device ofclaim 43, wherein the electrical characteristic is a potentialdifference across the membrane.
 48. The device of claim 43, wherein theelectrical characteristic is a short-circuit current condition acrossthe membrane.
 49. The device of claim 43, wherein the electricalcharacteristic is a resistance across the membrane